It is crucial to develop a miniaturized cultivation method for large and rapid screening of high-yielding mutants of monacolin-K, a powerful anti-hypercholesterolemic secondary metabolite biosynthesized by the fungal cells of Monascus ruber. In order to investigate as many strains as possible in a short time, a miniaturized fermentation method especially suitable for the cultivation of the filamentous Monascus mutants was developed using $50m{\ell}$ culture-tube ($7m{\ell}$ of working volume) instead of the traditional $250m{\ell}$ flask ($50m{\ell}$ of working volume). Generally, in filamentous fungal cell fermentations, morphologies in growth and production cultures should be maintained as thick filamentous and compact-pelleted (usually less than 1 mm in diameter) forms, respectively, for enhanced production of secondary metabolites in final production cultures. In this study, we intended to induce the respective optimal morphologies in the miniaturized culture system for the purpose of rapid screening of overproducers. Miniaturized growth culture system was successfully developed due to the mass production of spores in the statistically optimized solid medium. When large amounts of spores were inoculated into the growth cultures, and brown rice flour (20 g/L) was also supplemented to the growth medium, dense filamentous morphologies were successfully induced in the growth cultures performed with the 50 ml culture tubes. It was implied that the amounts of spores inoculated into the growth tube-cultures and the growth medium components should be the key factors for the induction of the filamentous forms in the growth fermentations. Furthermore, in order to statistically optimize production medium, multiple experiments based on Plackett-Burman design and response surface method (RSM) were carried out, resulting in more than 2 fold enhanced production of monacolin-K in the final production cultures with the optimized production medium. Notably, under the production culture conditions with the statistically optimized medium, optimal pellet sizes below 1 mm in diameter were reproducibly induced, in contrast to the thick and viscous filamentous morphologies observed in the previous production cultures.
For large and rapid screening of high-yielding mutants of lovastatin produced by filamentous fungal cells of Aspergillus terreus, one of the most important stage is to test as large amounts of mutated strains as possible. For this purpose, we intended to develop a miniaturized cultivation method using $7m{\ell}$ culture tube instead of traditional $250m{\ell}$ flask (working volume $50m{\ell}$). For obtaining large amounts of conidiospores to be used as inoculums for miniaturized cultures, 4 components i.e., glucose, sucrose, yeast extract and $KH_2PO_4$ were intensively investigated, which had been observed to show positive effect on enhancement of spore production through Plackett-Burman design experimet. When optimum concentrations of these components that were determined through application of response surface method (RSM) based on central composite design (CCD) were used, maximum spore numbers amounting to $1.9\times10^{10}$ spores/plate were obtained, resulting in approximately 190 fold increase as compared to the commonly used PDA sporulation medium. Using the miniaturized cultures, intensive strain development programs were carried out for screening of lovastatin high-yielding as well as highly reproducible mutants. It was observed that, for maximum production of lovastatin, the producers should be activated through 'PaB' adaptation process during the early solid culture stage. In addition, they should be proliferated in condensed filamentous forms in miniaturized growth cultures, so that optimum amounts of highly active cells could be transferred to the production culture-tube as reproducible inoculums. Under these highly controlled fermentation conditions, compact-pelleted morphology of optimum size (less than 1 mm in diameter) was successfully induced in the miniaturized production cultures, which proved essential for maximal utilization of the producers' physiology leading to significantly enhanced production of lovastatin. As a result of continuous screening in the miniaturized cultures, lovastatin production levels of the 81% of the daughter cells derived from the high-yielding producers turned out to be in the range of 80%$\sim$120% of the lovastatin production level of the parallel flask cultures. These results demonstrate that the miniaturized cultivation method developed in this study is efficient high throughput system for large and rapid screening of highly stable and productive strains.
Pleurotus ostreatus, the oyster mushroom, is one of the most widely cultivated and important edible mushrooms in the world. In order to study the developmental process of P. ostreatus and its regulatory mechanism, a new culturing method needs to be established for inducing the fruiting body and sporulation in the laboratory. In this study, we have examined whether the fruiting body of P. ostreatus can be formed on the plastic petri dish which are commonly used for cell culture in the laboratory. The strain was cultured on $60{\times}15mm$ plastic petri dish with potato dextrose agar media at $28^{\circ}C$ for mycelial growth and then at $18^{\circ}C$ for the formation of primordia and fruiting bodies within plant growth chamber. The development of primordia into fruiting bodies was achieved on cultured dishes under air ventilation. At the primordia stage, the normal formation of fruiting body was blocked by sealing the plastic dish with parafilm. The periods requiring for the formation of primordia and fruiting bodies were examined on the dish culture. About 96% and 76% of cultured samples formed primordia and fruiting bodies under the optimal conditions during ten weeks of culture, respectively. These culturing periods, however, were changed by the mechanical injury treatment to mycelia. As other factors affecting the fruiting body formation, the effects of light and cold shock have been tested. No fruiting formation was observed on the cultured dishes under the dark. The cold shock treatment by storing cultured dishes for one day at $4^{\circ}C$ did not have any significant effects in the fruiting body formation. Spores of fruiting bodies acquired from the petri dishes could be germinated on culture media at $28^{\circ}C$. These results suggest that the fruiting bodies of P. ostreatus can be formed on the experimental petri dish and this dish-culturing method is useful for understanding of the developmental process of P. ostreatus in the laboratory. Furthermore, the dish-culturing method is able to shorten the life cycle of P. ostreatus without requiring large area and expensive device.
Journal of the Korean Institute of Landscape Architecture
/
v.37
no.3
/
pp.54-60
/
2009
The purpose of this study is to analyze the impacts of three-dimensional land cover on changing urban air temperatures and to explore some strategies of urban landscaping towards mitigation of heat build-up. This study located study spaces within a diameter of 300m around 24 Automatic Weather Stations(AWS) in Seoul, and collected data of diverse variables which could affect summer energy budgets and air temperatures. The study also selected reflecting study objectives 6 smaller-scale spaces with a diameter of 30m in Chuncheon, and measured summer air temperatures and three-dimensional land cover to compare their relationships with results from Seoul's AWS. Linear regression models derived from data of Seoul's AWS revealed that vegetation volume, greenspace area, building volume, building area, population density, and pavement area contributed to a statistically significant change in summer air temperatures. Of these variables, vegetation and building volume indicated the highest accountability for total variability of changes in the air temperatures. Multiple regression models derived from combinations of the significant variables also showed that both vegetation and building volume generated a model with the best fitness. Based on this multiple regression model, a 10% increase of vegetation volume decreased the air temperatures by approximately 0.14%, while a 10% increase of building volume raised them by 0.26%. Relationships between Chuncheon's summer air temperatures and land cover distribution for the smaller-scale spaces also disclosed that the air temperatures were negatively correlated to vegetation volume and greenspace area, while they were positively correlated to hardscape area. Similarly to the case of Seoul's AWS, the air temperatures for the smaller-scale spaces decreased by 0.32% ($0.08^{\circ}C$) as vegetation volume increased by 10%, based on the most appropriate linear model. Thus, urban landscaping for the reduction of summer air temperatures requires strategies to improve vegetation volume and simultaneously to decrease building volume. For Seoul's AWS, the impact of building volume on changing the air temperatures was about 2 times greater than that of vegetation volume. Wall and rooftop greening for shading and evapotranspiration is suggested to control atmospheric heating by three-dimensional building surfaces, enlarging vegetation volume through multilayered plantings on soil surfaces.
Jang, Dae Gyu;Ahn, Hosang;Kim, Jeong Yeon;Ahn, Chang Hyuk;Lee, Saeromi;Kim, Jong Kyu;Joo, Jin Chul
Journal of Korean Society of Environmental Engineers
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v.36
no.11
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pp.771-780
/
2014
In this study, the various organic supports (i.e., silicone, acrylonitrile-butadiene-styrene, epoxy, and, butadiene rubber) with great sorption capacity of organic contaminants were chosen to develop nano-ZnO/organic composites (NZOCs) and to prevent the detachment of nano-ZnO particles. The water resistance of the developed NZOCs were evaluated, and the feasibility of the developed NZOCs were investigated by evaluating the removal efficiency of 1,1,2-trichloroethylene (TCE) in the aqueous phase. Based on the results from water-resistance experiments, long-term water treatment usage of all NZOCs was found to be feasible. According to the FE-SEM, EDX, and imaging analysis, nano-ZnO/butadiene rubber composite (NZBC) with various sizes and types of porosity and crack was measured to be coated with relatively homogeneously-distributed nano-ZnO particles whereas nano-ZnO/silicone composite (NZSC), nano-ZnO/ABS composite (NZAC), and nano-ZnO/epoxy composite (NZEC) with poorly-developed porosity and crack were measured to be coated with relatively heterogeneously-distributed nano-ZnO particles. The sorption capacity of NZBC was close to 60% relative to the initial concentration, and this result was mainly attributed to the amorphous structure of NZBC, hence the hydrophobic partitioning of TCE to the amorphous structure of NZBC intensively occurred. The removal efficiency of TCE in aqueous phase using NZBC was close to 99% relative to the initial concentration, and the removal efficiency of TCE was improved as the amount of NZBC increased. These results stemmed from the synergistic mechanisms with great sorption capability of butadiene rubber and superior photocatalytic activities of nano-ZnO. Finally, the removal efficiency of TCE in aqueous phase using NZBC was well represented by linear model ($R^2{\geq}0.936$), and the $K_{app}$ values of NZBC were from 2.64 to 3.85 times greater than those of $K_{photolysis}$, indicating that butadiene rubber was found to be the suitable organic supporting materials with enhanced sorption capacity and without inhibition of photocatalytic activities of nano-ZnO.
Kim, Chung-Soo;Kim, Eun-Tae;Lee, Jeong-Yong;Kim, Yong-Tae
Applied Microscopy
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v.38
no.4
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pp.279-284
/
2008
The phase change materials have been extensively used as an optical rewritable data storage media utilizing their phase change properties. Recently, the phase change materials have been spotlighted for the application of non-volatile memory device, such as the phase change random access memory. In this work, we have investigated the crystallization behavior and microstructure analysis of In-Sb-Te (IST) thin films deposited by RF magnetron sputtering. Transmission electron microscopy measurement was carried out after the annealing at $300^{\circ}C$, $350^{\circ}C$, $400^{\circ}C$ and $450^{\circ}C$ for 5 min. It was observed that InSb phases change into $In_3SbTe_2$ phases and InTe phases as the temperature increases. It was found that the thickness of thin films was decreased and the grain size was increased by the bright field transmission electron microscopy (BF TEM) images and the selected area electron diffraction (SAED) patterns. In a high resolution transmission electron microscopy (HRTEM) study, it shows that $350^{\circ}C$-annealed InSb phases have {111} facet because the surface energy of a {111} close-packed plane is the lowest in FCC crystals. When the film was heated up to $400^{\circ}C$, $In_3SbTe_2$ grains have coherent micro-twins with {111} mirror plane, and they are healed annealing at $450^{\circ}C$. From the HRTEM, InTe phase separation was occurred in this stage. It can be found that $In_3SbTe_2$ forms in the crystallization process as composition of the film near stoichiometric composition, while InTe phase separation may take place as the composition deviates from $In_3SbTe_2$.
Journal of the Korean Society of Food Science and Nutrition
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v.38
no.3
/
pp.384-390
/
2009
Extraction characteristics of citron (Citrus junos Sieb. ex Tanaka) seeds and functional properties of corresponding extract were monitored by response surface methodology (RSM). Maximum extraction yield of 20.23% was obtained at extraction temperature of $50.23^{\circ}C$, extraction time of 3.03 hr, and shaking velocity of 400.06 rpm. At extraction temperature, extraction time, and shaking velocity of $49.88^{\circ}C$, 2.72 hr, and 400.39 rpm, respectively, maximum polyphenol content was 4.37 mg/g. At extraction temperature, extraction time, and shaking velocity of $50.28^{\circ}C$, 3.42 hr, and 399.96 rpm, respectively, maximum electron donating ability (EDA) was 49.69%. Maximum nitrite scavenging activity (NSA) was 47.79% at extraction temperature, extraction time, and shaking velocity of $49.19^{\circ}C$, 0.68 hr, and 602.95 rpm, respectively. Based on superimposition of 3-dimensional RSM with respect to extraction yield, polyphenol, EDA, and NSA, optimum ranges of extraction conditions were extraction temperature of $50^{\circ}C$, extraction time of about 3 hr, and shaking velocity of 400 rpm.
Chlorophyll fluorescence, chlorophyll content, graft-taking and growth of grafted cucumber seedlings as affected by photosynthetic photon flux (PPF) of LED lamps were analyzed in this study. Four PPF levels, namely 25, 50, 100, $150{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$ were provided to investigate the effect of light intensity on the chlorophyll fluorescence, chlorophyll content, graft-taking and growth of grafted cucumber seedlings. Air temperature, relative humidity, and photoperiod for graft-taking were maintained at $25^{\circ}C$, 90%, $16h{\cdot}d^{-1}$, respectively. Maximum quantum yield (Fv/Fm) of rootstock as affected by PPF was found to be 0.84-0.85 and there was no significant change in Fv/Fm. Even though Fv/Fm of scion measured at 2 days after grafting was lowered to 0.81-0.82, after then it gradually increased with increasing PPF. At 4 days after grafting, the chlorophyll content extracted from scion increased with increasing PPF. Graft-taking ratio of grafted cucumber seedlings was 90-95% as PPF was ranged from $25{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$ to $100{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$. However, the graft-taking ratio of grafted seedlings healed under PPF of $150{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$ was decreased to 80%. Maximum PPF measured required for smooth joining of rootstock and scion was assumed to be $100{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$. At healing stage of grafted cucumber seedlings, Fv/Fm of scion decreased and at least two days after grafting were required for rooting of grafted seedlings. Chlorophyll fluorescence response of rootstock and scion was linked to light irradiation. Therefore, it was concluded that physical environment including light and humidity during healing process of grafted seedlings should be controlled more precisely to facilitate root formation and to prevent scion from lowering Fv/Fm. Further studies are required to investigate the effects of root development and joining of vascular bundles of grafted seedlings on the chlorophyll content of scion.
The Acorn (Quercus acutissima CARRUTHERS), which contains a large quantity of tannin, should be developed as a processed food as the acorn is rich in natural antioxidants and other valuable components. Accordingly, acorn extraction conditions for polyphenol and gallic acid (both antioxidants) were investigated by response surface methodology. The content of polyphenols were determined under 16 different extraction conditions based upon a central composite design. The parameters varied over $30-70^{\circ}C$ of extraction temperature, 1-5 h of extraction time, and 5-25 mL/g of solvent ratio, Gallic acid extraction was optimal at $60-100^{\circ}C$ extraction temperature, 1-5 h of extraction time, and 5-25 mL/g of solvent ratio, Epicatechin content was highest at $56.77^{\circ}C$, 4.16 hand 22.38 mL/g. Catechin content was highest at $52.37^{\circ}C$, 2h and 23.59 mL/g. The maximum catechin content was $91.30{\mu}g/mL$. Epigallocatechin content was influenced by extraction temperature and time. The maximum epigallocatechin content was $1,066.56{\mu}g/mL$ at $61.42^{\circ}C$, 4.17h, and 9.25 mL/g. The maximum value of epicatechingallate content was $125.39{\mu}g/mL$ at $47.72^{\circ}C$, 3.04h, and 24.93mL/g. Epigallocatechingallate content was influenced principally by solvent ratio and the maximum content was $61.38{\mu}g/mL$ at $48.11^{\circ}C$, 2.96h, and 24.95mL/g. The total polyphenol content was maximal at $1,332.75{\mu}g/mL$, after extraction at $61.50^{\circ}C$, 4.24h, at 9.71mL/g. The higher the extraction temperature and the longer the extraction time, the greater the polyphenol content. Gallic acid content was highest, the maximal level was $30.51{\mu}g/mL$ after $65.84^{\circ}C$, 1.65h at 17.17 mL/g, and this was influenced principally by extraction time and solvent ratio.
This study was carried out to reduce the loss of frozen dough quality during frozen storage. Using response surface method, ascorbic acid 160.4 ppm, L-cysteine 63.1 ppm, and SSL 0.6% were found to be optimum, with xanthan gum 0.3% (formula A) and Ultra tex-3 5% (formula B) added as cryoprotectants. During frozen storage at $-20^{\circ}C$, control rapidly deteriorated after 4 weeks, while formulas A and B showed slight deterioration with immutable quality after 10 weeks.
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