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Integrated Rotary Genetic Analysis Microsystem for Influenza A Virus Detection

  • Jung, Jae Hwan;Park, Byung Hyun;Choi, Seok Jin;Seo, Tae Seok
    • Proceedings of the Korean Vacuum Society Conference
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    • 2013.08a
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    • pp.88-89
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    • 2013
  • A variety of influenza A viruses from animal hosts are continuously prevalent throughout the world which cause human epidemics resulting millions of human infections and enormous industrial and economic damages. Thus, early diagnosis of such pathogen is of paramount importance for biomedical examination and public healthcare screening. To approach this issue, here we propose a fully integrated Rotary genetic analysis system, called Rotary Genetic Analyzer, for on-site detection of influenza A viruses with high speed. The Rotary Genetic Analyzer is made up of four parts including a disposable microchip, a servo motor for precise and high rate spinning of the chip, thermal blocks for temperature control, and a miniaturized optical fluorescence detector as shown Fig. 1. A thermal block made from duralumin is integrated with a film heater at the bottom and a resistance temperature detector (RTD) in the middle. For the efficient performance of RT-PCR, three thermal blocks are placed on the Rotary stage and the temperature of each block is corresponded to the thermal cycling, namely $95^{\circ}C$ (denature), $58^{\circ}C$ (annealing), and $72^{\circ}C$ (extension). Rotary RT-PCR was performed to amplify the target gene which was monitored by an optical fluorescent detector above the extension block. A disposable microdevice (10 cm diameter) consists of a solid-phase extraction based sample pretreatment unit, bead chamber, and 4 ${\mu}L$ of the PCR chamber as shown Fig. 2. The microchip is fabricated using a patterned polycarbonate (PC) sheet with 1 mm thickness and a PC film with 130 ${\mu}m$ thickness, which layers are thermally bonded at $138^{\circ}C$ using acetone vapour. Silicatreated microglass beads with 150~212 ${\mu}L$ diameter are introduced into the sample pretreatment chambers and held in place by weir structure for construction of solid-phase extraction system. Fig. 3 shows strobed images of sequential loading of three samples. Three samples were loaded into the reservoir simultaneously (Fig. 3A), then the influenza A H3N2 viral RNA sample was loaded at 5000 RPM for 10 sec (Fig. 3B). Washing buffer was followed at 5000 RPM for 5 min (Fig. 3C), and angular frequency was decreased to 100 RPM for siphon priming of PCR cocktail to the channel as shown in Figure 3D. Finally the PCR cocktail was loaded to the bead chamber at 2000 RPM for 10 sec, and then RPM was increased up to 5000 RPM for 1 min to obtain the as much as PCR cocktail containing the RNA template (Fig. 3E). In this system, the wastes from RNA samples and washing buffer were transported to the waste chamber, which is fully filled to the chamber with precise optimization. Then, the PCR cocktail was able to transport to the PCR chamber. Fig. 3F shows the final image of the sample pretreatment. PCR cocktail containing RNA template is successfully isolated from waste. To detect the influenza A H3N2 virus, the purified RNA with PCR cocktail in the PCR chamber was amplified by using performed the RNA capture on the proposed microdevice. The fluorescence images were described in Figure 4A at the 0, 40 cycles. The fluorescence signal (40 cycle) was drastically increased confirming the influenza A H3N2 virus. The real-time profiles were successfully obtained using the optical fluorescence detector as shown in Figure 4B. The Rotary PCR and off-chip PCR were compared with same amount of influenza A H3N2 virus. The Ct value of Rotary PCR was smaller than the off-chip PCR without contamination. The whole process of the sample pretreatment and RT-PCR could be accomplished in 30 min on the fully integrated Rotary Genetic Analyzer system. We have demonstrated a fully integrated and portable Rotary Genetic Analyzer for detection of the gene expression of influenza A virus, which has 'Sample-in-answer-out' capability including sample pretreatment, rotary amplification, and optical detection. Target gene amplification was real-time monitored using the integrated Rotary Genetic Analyzer system.

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Introduction of Kjeldahl Digestion Method for Nitrogen Stable Isotope Analysis (δ15N-NO3 and δ15NNH4) and Case Study for Tracing Nitrogen Source (Kjeldahl 증류법을 활용한 질산성-질소 및 암모니아성-질소 안정동위원소비 분석 및 질소오염원 추적 사례 연구)

  • Kim, Min-Seob;Park, Tae-Jin;Yoon, Suk-Hee;Lim, Bo-La;Shin, Kyung-Hoon;Kwon, Oh-Sang;Lee, Won-Seok
    • Korean Journal of Ecology and Environment
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    • v.48 no.3
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    • pp.147-152
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    • 2015
  • Nitrogen (N) loading from domestic, agricultural and industrial sources can lead to excessive growth of macrophytes or phytoplankton in aquatic environment. Many studies have used nitrogen stable isotope ratios to identify anthropogenic nitrogen in aquatic systems as a useful method for studying nitrogen cycle. In this study to evaluate the precision and accuracy of Kjeldahl processes, two reference materials (IAEA-NO-3, N-1) were analyzed repeatedly. Measured the ${\delta}^{15}N-NO_3$ and ${\delta}^{15}N-NH_4$ values of IAEA-NO-3 and IAEA-N-1 were $4.7{\pm}0.2$‰ and $0.4{\pm}0.3$‰, respectively, which are within recommended values of analytical uncertainties. Also, we investigated spatial patterns of ${\delta}^{15}N-NO_3$ and ${\delta}^{15}N-NH_4$ in effluent plumes from a waste water treatment plant in Han River, Korea. ${\delta}^{15}N-NO_3$ and ${\delta}^{15}N-NH_4$ values are enriched at downstream areas of water treatment plant suggesting that dissolved nitrogen in effluent plumes should be one of the main N sources in those areas. The current study clarifies the reliability of Kjeldahl analytical method and the usefulness of stable isotopic techniques to trace the contamination source of dissolved nitrogen such as nitrate and ammonia.

Bactericidal Efficacy of a Fumigation Disinfectant with Ortho-phenylphenol as an Active Ingredient Against Pseudomonas Aeruginosa and Enterococcus Hirae (Ortho-phenylphenol을 주성분을 하는 훈증소독제의 Pseudomonas aeruginosa와 Enterococcus hirae에 대한 살균효과)

  • Cha, Chun-Nam;Park, Eun-Kee;Kim, Yongpal;Yu, Eun-Ah;Yoo, Chang-Yeol;Hong, Il-Hwa;Kim, Suk;Lee, Hu-Jang
    • Journal of Food Hygiene and Safety
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    • v.29 no.1
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    • pp.60-66
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    • 2014
  • This test was performed to evaluate the bactericidal efficacy of a fumigation disinfectant containing 20% ortho-phenylphenol against Pseudomonas aeruginosa (P. aeruginosa) and Enterococcus hirae (E. hirae). In preliminary tests, P. aeruginosa and E. hirae working culture suspension number (N value) were $2.8{\times}10^8$ and $4.0{\times}10^8CFU/mL$, respectively. And all the colony numbers on the carriers exposed to the fumigant (n1, n2, n3) were higher than 0.5N1 (the number of bacterial test suspentions by pour plate method), 0.5N2 (the number of bacterial test suspentions by filter membrane method) and 0.5N1, respectively. In addition, the mean number of P. aeruginosa and E. hirae recovered on the control-carriers (T value) was $2.8{\times}10^8$ and $3.4{\times}10^6CFU/mL$, respectively. In the bactericidal effect of the fumigant, the reduction number of $2.8{\times}10^8$ (d value) was 6.46 and 5.19 logCFU/mL, respectively. According to the French standard for the fumigant, the d value for the effective bactericidal fumigant should be over than 5 logCFU/mL. With the results from this study, the fumigation disinfectant containing 20% ortho-phenylphenol has an effective bactericidal activity, then the fumigant can be applied to disinfect food materials and kitchen appliances contaminated with the pathogenic bacteria.

Effect of Pre-Fermentation Condition on Quality Characteristics of Small Packaged Cabbage Kimchi (소포장 양배추김치의 품질특성에 미치는 전 발효조건의 영향)

  • Kim, Yoo-Won;Han, Seo-Young;Choi, Hye-Sun;Han, Gwi-Jung;Park, Hye-Young
    • Korean journal of food and cookery science
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    • v.28 no.4
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    • pp.391-397
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    • 2012
  • This study was carried out to investigate commercialization of Kimchi made of cabbage (Brassica oleracea var. capitata L.) using pre-fermentation conditions. The pre-fermentation conditions were 0, 18, 24, and 28 h at $20^{\circ}C$, and then the samples were stored at $10^{\circ}C$ to assess changes in quality characteristics. A comparison of the quality characteristics during storage showed that PF24 (pre-fermented cabbage Kimchi during 24 h at $20^{\circ}C$) and PF28 (pre-fermented cabbage Kimchi during 28 h at $20^{\circ}C$) had pH 4.47 and pH 4.23 on the second day of storage, respectively. It was possible to shorten the fermentation time to less than that of PF0 (not pre-fermented cabbage Kimchi at $20^{\circ}C$), by approximately 3 days. Total acidity was 0.26 to 0.29% immediately after making the Kimchi. However, PF0, PF18 (pre-fermented cabbage Kimchi during 18 h at $20^{\circ}C$), PF24 and PF28 became well-fermented when they were stored for 8~14 days, 3~10 days or 2~3 days. The number of lactic acid bacteria increased with the passage of time in all treatment groups regardless of fermentation conditions. However, the longer pre-fermentation time became, the faster the number of lactic acid bacteria increased. Most samples showed similar results late in the storage period; 7.2~7.4 log CFU/mL. PF0 had the greatest volume change 2.1 times increase late in the storage period. The sensory evaluation showed significant differences for flavor, taste, and overall acceptability after a partial storage period. PF28 stored for 2~3 days showed excellent flavor, and PF24 and PF28 stored for 2~3 days showed the highest scores of 6.27 to 6.67. The PF24 and PF28 treated samples were appropriate for commercializing small packed cabbage Kimchi and for alleviating the expansion problem of the packing material. However, because mass commercial production requires a large number of samples to be used at once, the results should be assessed for industrial product development in the future.

Antioxidant Activity and Quality Characteristics of Soy Milk Mayonnaise Containing Soybean Paste Powder (된장분말을 첨가한 두유마요네즈의 항산화성 및 품질특성)

  • Park, Hye Duck;Lee, Sang Sun
    • Food Engineering Progress
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    • v.13 no.3
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    • pp.203-210
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    • 2009
  • In this study, antioxidant activity and quality characteristics of egg yolk mayonnaise and soy milk mayonnaise containing soybean paste powder were assessed. The mayonnaise with egg yolk as an emulsifier is E group according to the amount of soybean paste powder addition of 0% (E1), 3% (E2), 6% (E3) and mayonnaise with soy milk as an emulsifier is S group with soybean paste powder addition of 0% (S1), 3% (S2), 6% (S3). Total phenol content and electron donating ability of mayonnaise were increased as the amount of the soybean paste powder increased. L value of Hunter's color was decreased, but a value and b value were increased as the amount of the soybean paste powder increased. Soy milk mayonnaise showed very low cholesterol content than egg yolk mayonnaise. In sensory evaluation, S2 received the highest scores for flavor, overall taste and preference. Base on these results, soybean paste powder showed significant antioxidant activity in mayonnaise and cholesterol content was successfully decreased in soy milk mayonnaise compared to egg yolk mayonnaise. The soy milk mayonnaise with soybean paste powder will be a possible model to introduce our traditional food to the worldwide food item, such as mayonnaise.

Detection of Spoilage Odors in Beef Using R-index and Pseudomonas Growth during Storage (쇠고기의 저장 중 R-index에 의한 부패취 발생시점과 Pseudomonas의 증식과의 비교 분석)

  • Byeon, Ko Eun;Park, Han Jo;An, Soo Rim;Hong, Kwang Won;Min, Sang Gi;Chung, Ku Young;Won, Kee Hoon;Lee, Seung Ju
    • Food Engineering Progress
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    • v.13 no.2
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    • pp.117-121
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    • 2009
  • Detection of spoilage odors from beef during storage was investigated using sensory evaluation with R-index, and microbial assay for Pseudomonas. Beef samples were tested to measure the flavor changes, which were converted to R-index, and the Pseudomonas levels during storage. There was a steep rise in R-index until 36 hr after storage at 25$^{\circ}C$, and then a gentle rise from 48 hr, whereas, there was a steady rise in R-index in the whole range of storage at 5$^{\circ}C$. Detection time of spoilage odors according to R-index was statistically analyzed at $\alpha$=5% to be at 30.92${\pm}$3.47 hr and 169.80${\pm}$11.27 hr for 25 and 5$^{\circ}C$ storage, respectively, and analyzed at $\alpha$=1% to be 34.80${\pm}$4.01 and 176.41${\pm}$9.89 hr for 25 and 5$^{\circ}C$ storage, respectively. At the detection times of spoilage odors, the Pseudomonas levels were found to be almost the same, but less than 6-7 log CFU/g generally known as a standard level at occurrence of spoilage odors in beef. This indicated that some other factors than the Pseudomonas reactions could be associated with generation of spoilage odors.

Effect of Green Tea and Saw Dust Contents on Static Bending Strength Performance of Hybrid Boards Composed of Wood Fiber, Saw Dust and Green Tea (목재섬유, 톱밥 및 녹차 이종복합보드의 정적 휨 강도성능에 미치는 녹차 및 톱밥 배합비율의 영향)

  • Park, Han-Min;Lee, Soo-Kyeong;Seok, Ji-Hoon;Choi, Nam-Kyung;Kwon, Chang-Bae;Heo, Hwang-Sun;Kim, Jong-Chul
    • Journal of agriculture & life science
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    • v.45 no.6
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    • pp.41-46
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    • 2011
  • In this study, in addition to the green tea-wood fiber hybrid composite boards of previous researches, to make effective use of saw dust of domestic cypress tree with functionalities and application as interior materials, eco-friendly hybrid composite boards were manufactured from wood fiber, green tea and saw dust of cypress tree. We investigated the effect of the component ratio of saw dust and green tea on static bending strength performances. Static bending MOE (modulus of elasticity) was within 0.956~1.18GPa, and showed the highest value in wood fiber : green tea : saw dust = 50 : 40 : 10 of the component ratio, and had the lowest value in 50 : 30 : 20 of component ratio. These values were 2.0~3.1times lower than those of green tea-wood fiber hybrid composite boards reported in the previous researches. The bending MOR (modulus of rupture) showed 8.99~11.5MPa, the change of the bending MOR with component ratio of the factors was the same as that of bending MOE. These values had 1.9~3.5 times lower value than those of green tea-wood fiber hybrid composite boards, and showed the slightly lower values than the MOR of particle boards (PB) and medium density fiberboards (MDF) prescribed in Korean Industrial Standard. Therefore, it is considered that these hybrid composite boards need to improve strength performances by component ratio change, hybrid composite with other materials and adhesive change etc. in order to industrialize the hybrid composite boards.

Effect of Different Levels of Applications of Illite on the Growth of Red Pepper (Capsicum annuum L.) in Bed Soil (상토에서 일라이트의 혼합비율에 따른 고추 육묘시 생육효과)

  • Lee, Seok-Eon;Kim, Hong-Ki;Kwon, Sang-Moon;Kim, Hee-Jung;Yoo, Ri-Bi;Baek, Ki-Tae;Lee, Moon-Soon;Woo, Sun-Hee;Park, Man;Chung, Keun-Yook
    • Korean Journal of Soil Science and Fertilizer
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    • v.43 no.6
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    • pp.852-857
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    • 2010
  • This study was performed to explore the effect of the clay mineral illite on the improvement of bed soil and plant growth. Red pepper (Capsicum annuum L.) was used as a model vegetable crop. The experiment was performed during the whole six weeks in the glass house of the Chungbuk National University. Its seedlings were cultivated in the bed soil normally used for horticultural purpose. Of the seedlings cultured, the healthy and regular size of seed were selected and cultivated in the pots. They were treated with two forms of illite, particulate (PA) and powder (PW), at the following application rates: standard application[P1 (PA1, PW1), 1:20 (w/w)], two times[P2 (PA2, PW2), 1:10 (w/w)], and four times[P4 (PA4, PW4), 1:5 (w/w)] of standard application. Untreatment (P0) was used as a control pot. At six weeks of cultivation, their growth lengths were correspondingly increased as the application rate was increased ranging from P0, P1, P2, and to P4. Their growth length was a little greater on the application of powder illite (PW) than on the particulate illite (PA). Based on the plant analysis for the root, leaf, stem of red pepper, the uptake amounts of K, Ca, and Mg, were correspondingly increased, as the application rate was increased ranging from P0, P1, P2, and to P4, respectively. At the same application rate, their amounts taken up in the respective parts were higher on the application of PW illite than on the PA one. Especially the amounts of Ca and Mg were higher in the stem, leaf than root. Consequently, it appears that the illite treatment, especially, PW form of illite, enhance the growth of red pepper in the glass house during the whole six weeks of experiment.

Influences of Chinese Cabbage Growth and Soil Salinity to Alternative Irrigation Waters (대체관개 용수에 의한 배추생육 및 토양 염류도에 미치는 영향)

  • Shin, Joung-Du;Park, Sang-Won;Kim, Won-Il;Lee, Jong-Sik;Yun, Sun-Gang;Eom, Ki-Cheol
    • Korean Journal of Soil Science and Fertilizer
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    • v.40 no.1
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    • pp.25-30
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    • 2007
  • Objective of this experiment was to investigate the growth effects of Chinese cabbage and soil salinity to alternative irrigation waters for drought periods. The treatments were consisted of the discharge water from industrial wastewater treatment plant (DIWT), the discharge water from municipal wastewater treatment plant (DMWT) and ground water as the control. For the chemical compositions of alternative water, it appeared that concentrations of the $Ni^+$ and SAR values in DIWT were over the reuse criteria of other countries for irrigation, but CODcr concentration in DMWT was higher than the reuse criteria for agricultural irrigation. According to classification of water by $EC_i$ value, DIWT and DMWT are ranged from 0.7 to $2.0dS\;m^{-1}$, slight salinity. Average harvest indexes were 0.64 for DIWT and 0.63 for DMWT as compared to 0.61 of the control regardless of irrigation periods. SAR value in soil was increased with prolonging the irrigation periods at head forming stage, but not much difference except for 30 days of irrigation period at harvesting time for DIWT. However, it was not much difference along with irrigation periods through the growth stages for DMWT as compared with the groundwater. At harvesting time, average $EC_e$ for the soil irrigated with alternative agricultural waters was $0.017dS\;m^{-1}$ for its DIMT and $0.036dS\;m^{-1}$ for its DMWT as compared to $0.013dS\;m^{-1}$ of its groundwater as the control. For $NH_4-N$ concentrations, it observed that there were no differences among the treatments with different irrigation periods at head forming stage in soil after irrigation. Also, $NO_3-N$ concentration in soil was increased up to 20 days after irrigation, and then decreased at 30 days after irrigation with DMWT at head forming stage. The $Ni^+$ concentration in upper layer soil (0-15 cm) irrigated with DIWT was increased with prolonging the irrigation period at head forming stage, but it was dramatically decreased and almost constant in all the treatments at harvesting time. Therefore, it might be concluded that there was potentially safe to irrigate the discharge water from municipal wastewater treatment plant for 20 days after transplanting to drought periods with cultivating the Chinese cabbage.

Comparison on the Extraction Efficiency and Antioxidant Activity of Flavonoid from Citrus Peel by Different Extraction Methods (추출방법에 따른 감귤 과피 유래 Flavonoid의 추출효율 및 항산화 효과에 대한 비교)

  • Cheigh, Chan-Ick;Jung, Won-Guen;Chung, Eun-Young;Ko, Min-Jung;Cho, Sang-Woo;Lee, Jae-Hwan;Chang, Pahn-Shick;Park, Young-Seo;Paik, Hyun-Dong;Kim, Kee-Tae;Chung, Myong-Soo
    • Food Engineering Progress
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    • v.14 no.2
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    • pp.166-172
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    • 2010
  • The extraction of polyphenol and flavonoid from citrus peel was performed by the ethanol, sugar, hot water (80$^{\circ}C$), and subcritical water extraction methods. The maximum yields of total polyphenolic compounds (27.25${\pm}$1.33 mg QE/g DCP, QE and DCP indicate quercetin equivalent and dried citrus peel, respectively) and flavonoids (7.31${\pm}$0.41 mg QE/g DCP) were obtained by subcritical water extraction (SWE) with operating conditions of 190$^{\circ}C$, 1300 psi, and 10 min. The yields by SWE were over 7.2-, and 8.5-fold higher than those of total polyphenols (3.79${\pm}$0.73 mg QE/g DCP) and flavonoids (0.86${\pm}$0.27 mg QE/g DCP) obtained using the ethanol extraction, which showed the highest extraction efficiency among tested conventional methods, respectively. Antioxidant activities of extracts obtained by different methods showed no significant differences. However, the relative antioxidant yield per 1 g dried citrus peel by SWE (190$^{\circ}C$, 10 min) was over 9.5-fold higher than that by the ethanol extraction.