• Title/Summary/Keyword: Separation control

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Optimization of SNP Genotyping Assay with Fluorescence Polarization Detection

  • Cai Chun Mei;Van Kyujung;Kim Moon Young;Lee Suk-Ha
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.50 no.5
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    • pp.361-367
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    • 2005
  • Single nucleotide polymorphisms (SNPs) are valuable DNA markers due to their abundance and potential for use in automated high-throughput genotyping. Numerous SNP genotyping assays have been developed. In this report, one of effective and high throughput SNP genotyping assays, which was named the template-directed dye-terminator incorporation with fluorescence polarization detection (FP-TDI) was described. Although the most of this assay succeed, the objective of this work was to deter­mine the reasons for the failures, find ways to improve the assay and reduce the running cost. Ninety $F_2$-derived soybean, Glycine max (L.) Merr., RILs from a cross between 'Pureunkong' and 'Jinpumkong 2' were genotyped at four SNPs. FP measurement was done on $Victot^3$ microplate reader (perkinelmer Inc., Boston, MA, USA). Increasing the number of thermal cycles in the single-base extension step increased the separation of the FP values between the products corresponding to different genotypes. But in some assays, excess of heterozygous genotypes was observed with increase of PCR cycles. We discovered that the excess heterozygous was due to misincorporation of one of the dye­terminators during the primer extension reaction. After pyrophosphatase incubation and thermal cycle control, misincoporation can be effectively prevented. Using long amplicons instead of short amplicons for SNP genotyping and decreasing the amount of dye terminator and Acyclopol Taq polymerase to 1/2 or 1/3 decreased the cost of the assay. With these minor adjustments, the FP-TDI assay can be used more accurately and cost-effectively.

Purification and Structural Analysis of Surfactin Produced by Endophytic Bacillus subtilis EBS05 and its Antagonistic Activity Against Rhizoctonia cerealis

  • Wen, Cai-Yi;Yin, Zhi-Gang;Wang, Kai-Xuan;Chen, Jian-Guang;Shen, Shun-Shan
    • The Plant Pathology Journal
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    • v.27 no.4
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    • pp.342-348
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    • 2011
  • Bacillus subtilis EBS05, an endophytic bacteria strain isolated from a medicinal plant Cinnamomum camphor, can produce antagonistic compounds that effectively inhibit plant pathogenic fungi. The greenhouse experiments showed that wheat sharp eyespot disease (WSED) was reduced by 91.2%, 88.2% and 43.0% after the treatment with fermentation broth, bacteria-free filter and a fungicide fludioxonil, respectively. The culture broth of strain EBS05 can more effectively control WSED than can fludioxonil. The fermentation broth and bacteria-free filter ability to suppress WSED was not significantly different, suggesting that an active secreted substance played a major role in controlling WSED. Separation and purification of the active compounds was carried out by serial processes, including hydrochloric acid (pH 2.0) treatment, methanol extraction and Sephadex LH-20 column chromatography, silica gel column chromatography and reverse-phase high-pressure liquid chromatography (HPLC), respectively. The purified compounds, one of active peaks in the HPLC spectrum, were obtained from the collection. Analysis of the chemical structures by time-of-flight mass spectrometry (TOF-MS) and electrospray ionization mass spectrometry/mass spectrometry (ESI-MS/MS) showed that the active substances produced by the endophytic bacteria EBS05 are mixture of the ${\beta}$-hydroxy-C12~C15-$Leu^7$ surfactin A isomers with 1035.65 Da, 1021.64 Da, 1007.63 Da and 993.65 Da molecular weights, respectively.

Carboxylic Acids as Biomarkers of Biomphalaria alexandrina Snails Infected with Schistosoma mansoni

  • Abou Elseoud, Salwa M. F.;Fattah, Nashwa S. Abdel;El Din, Hayam M. Ezz;Abdel Al, Hala;Mossalem, Hanan;Elleboudy, Noha
    • Parasites, Hosts and Diseases
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    • v.48 no.2
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    • pp.127-132
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    • 2010
  • Biomphalaria alexandrina snails play an indispensable role in transmission of schistosomiasis. Infection rates in field populations of snails are routinely determined by cercarial shedding neglecting prepatent snail infections, because of lack of a suitable method for diagnosis. The present study aimed at separation and quantification of oxalic, malic, acetic, pyruvic, and fumaric acids using ion-suppression reversed-phase high performance liquid chromatography (HPLC) to test the potentiality of these acids to be used as diagnostic and therapeutic biomarkers. The assay was done in both hemolymph and digestive gland-gonad complex (DGG) samples in a total of 300 B. alexandrina snails. All of the studied acids in both the hemolymph and tissue samples except for the fumaric acid in hemolymph appeared to be good diagnostic biomarkers as they provide not only a good discrimination between the infected snails from the control but also between the studied stages of infection from each other. The most sensitive discriminating acid was malic acid in hemolymph samples as it showed the highest F-ratio. Using the Z-score, malic acid was found to be a good potential therapeutic biomarker in the prepatency stage, oxalic acid and acetic acid in the stage of patency, and malic acid and acetic acid at 2 weeks after patency. Quantification of carboxylic acids, using HPLC strategy, was fast, easy, and accurate in prediction of infected and uninfected snails and possibly to detect the stage of infection. It seems also useful for detection of the most suitable acids to be used as drug targets.

Effect of Ethanol on Prostaglandins Production of Monocytes (에탄올이 단핵구의 Prostaglandins 생산에 미치는 영향)

  • 박란숙
    • Journal of Nutrition and Health
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    • v.24 no.2
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    • pp.97-103
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    • 1991
  • The increase in alcohol consumption level has been noticed in Korea recently. Alcohol appreciably inhibits cell mediated immunity and this may contribute to the high prevalence of serious infection such as pulmonary tuberculosis among alcoholic subjects. The present study was undertaken to examine the effect of ethanol on the cyclooxygenase metabolites of human monocyte in vitro. Monocytes were activated with 800 units of gamma interferon(IFN-${\gamma}$) for 3 days following apply of Ficool-hypaque density gradient and gelatin coated flasks for separation of monocytes. Ethanol with addition of 100mM, 300mM and 600 mM for 30 minutes to 106 monocytes with/without previous IFN-${\gamma}$ treatment caused a dose dependent decrease in the production of thromboxane B2, 6-keto-PGE1$\alpha$ and PGE2 by radioimmunoassay at 6 hours after ethanol treatment. Quite different from the findings after 6 hours there was dose dependent increase in three prostaglandins without IFN-${\gamma}$ treatment after 24 hours of incubation. With previous treatment of IFN-${\gamma}$ reduced productions of three prostaglandins at 24 hours than control is spite of ethanol stimjulation. These findings show that IFN-${\gamma}$ can inhibit alcohol induced derangement of arachidonic acid metabolism of monocytes.

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A Study on Cloud Network and Security System Analysis for Enhanced Security of Legislative Authority (입법기관의 보안강화를 위한 Cloud 네트워크 분석 및 보안 시스템 연구)

  • Nam, Won-Hee;Park, Dea-Woo
    • Journal of the Korea Institute of Information and Communication Engineering
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    • v.15 no.6
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    • pp.1320-1326
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    • 2011
  • National institutions on the importance of information security is being recognized, information security laws are being discussed in Congress 3.4 DDoS incident and Nonghyup hacking, etc. However, National Assembly Secretariat when the results of the Information Security Consulting has been assessed very low 61.2 points, evaluation of hardware and software in secure areas were vulnerable. This paper, the legislative support agencies National Assembly and National Assembly Secretariat on the network and computer systems, and managerial, technical and physical security elements are analyzed for the status. And network should have the legislative support agencies and system for the physical network separation, DDoS attack response, Virus attack response, hacking attacks response, and Cyber Emergency Response Team/Coordination Center for Cyber infringing design and research through the confidentiality, integrity, availability, access control, authentication and security analysis is based on the evaluation criteria. Through this study, the legislative support agencies to strengthen the security of data and security laws enacted to provide the basis for.

An Transport Layer Vertical Handover Approach for Video Services in Overlay Network Environments (오버레이 네트워크 환경에서 비디오 서비스를 위한 트랜스포트 계층에서의 수직 핸드오버 방안)

  • Chang, Moon-Jeong;Lee, Mee-Jeong
    • The KIPS Transactions:PartC
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    • v.14C no.2
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    • pp.163-170
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    • 2007
  • The next generation communication environment consists of various wireless access networks with distinct features that are configured as an overlay topology. In the network environments, the frequency of hand overs should be minimized and the error propagation should be solved in order to provide high-quality multimedia services to mobile users. Therefore, we propose an performance enhancement approach, based on mSCTP, that provides high quality multimedia services to mobile users by ameliorating the error propagation problem. We utilizes the following four functions: 1) the separation of transmission paths according to the types of frames. 2) retransmission strategy to minimize the loss rate of frames, 3) Foced vertical handover execution by utilizing bicasting, 4) using the stability period in order to reduce the effect of the ping pong phenomenon. The simulation results show that the proposed approach provides seamless multimedia service to mobile users by achieving error resilience.

Simultaneous Quantification of Three Marker Compounds in Samultang by HPLC/DAD (HPLC-DAD를 이용한 사물탕 중 3종 성분의 동시분석법 확립)

  • Won, Jin-Bae;Ma, Jin-Yeul;Lee, Jae-Hoon;Ma, Choong-Je
    • Korean Journal of Pharmacognosy
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    • v.40 no.4
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    • pp.298-302
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    • 2009
  • Samultang is one of traditional medicine composed of Paeonia lactiflora, Angelica gigas, Rehmannia glutinosa and Cnidium officinale. To develop simultaneous determination of paeoniflorin, decursin and 5-HMF in Samultang, a high performance liquid chromatography with diode array detector was used. To separate three marker components, Dionex $C_{18}$ column (5 ${\mu}m$, 120 ${\AA}$, 4.6 mm${\times}$150 mm) was used with a gradient elution system of water and methanol. UV wavelength of detector set at 230 nm and 280 nm. This method was validated by linearity, precision test and recovery test. Calibration curves of three standard components were showed good linear regression ($R^2$>0.9973). LOD and LOQ ranged from 0.08 ${\mu}g$/ml to 0.38 ${\mu}g$/ml and 0.25 ${\mu}g$/ml to 1.16 ${\mu}g$/ml, respectively. The relative standard deviations (RSDs) of data of the inter-day and intra-day experiments were less than 0.54% and 0.89%, respectively. The measured results of recovery test were varied from 93.36 to 107.79 with RSD values 0.01~1.45%. The established method was applied for separation of bio-conversion Samultang sample and compared with control sample.

Quantitative Analysis of Ellagic Acid in Nymphaea tetragona Georgi Root by HPLC-UVD (HPLC-UVD를 이용한 수련(Nymphaea tetragona Georgi)의 Ellagic Acid 정량 분석)

  • Lee, Dong Young;Jeong, Eun Ju;Jeon, Byung Ju;Sung, Sang Hyun
    • Korean Journal of Pharmacognosy
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    • v.45 no.1
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    • pp.84-87
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    • 2014
  • A high performance liquid chromatography (HPLC) method for the quantitation of ellagic acid in Nymphaea tetragona was developed for the quality control of functional cosmetic ingredient, the extract of N. tetragona. Separation and quantitation were successfully achieved with a Kromasil C18 column ($5{\mu}m$, $250mm{\times}4.6mm$, i.d.) by isocratic elution of a mixture of acetonitrile containing 0.1% trifluoroacetic acid and water containing 0.03% phosphoric acid at a flow rate of 1.0 ml/min. The UV detector was used for the detection and the wavelength for quantitation was set at 254 nm. The presence of ellagic acid in the extract was determined by comparison of retention time and spiking with authentic standard. Analytical results showed good linearity ($R^2=0.99996$) in relatively wide concentration ranges. The R.S.D. for precision test was less than 3.0%. Recovery of the compound was 98.55~101.72% with R.S.D values less than 4.0%. In conclusion, this method has been successfully applied to the determination of ellagic acid in N. tetragona.

Anti-adipogenic Pregnane Steroid from a Hydractinia-associated Fungus, Cladosporium sphaerospermum SW67

  • Lee, Seoung Rak;Kang, Heesun;Yoo, Min Jeong;Yu, Jae Sik;Lee, Seulah;Yi, Sang Ah;Beemelmanns, Christine;Lee, Jaecheol;Kim, Ki Hyun
    • Natural Product Sciences
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    • v.26 no.3
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    • pp.230-235
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    • 2020
  • A pregnane steroid, 3α-hydroxy-pregn-7-ene-6,20-dione (1), was isolated from a Hydractinia-associated Cladosporium sphaerospermum SW67 by repetitive column chromatographic separation and high-performance liquid chromatography (HPLC) purification. The planar structure of 1 was elucidated from the analysis of the spectroscopic data (1D and 2D NMR spectra) and LC-MS data. The absolute configuration of 1 was determined by interpretation of ROESY spectrum of 1, together with the comparison of reported spectroscopic values in previous studies. To the best of our knowledge, this is the first report of the identification of the pregnane scaffold from C. sphaerospermum, a natural source. Compound 1 was evaluated for its effects on lipid metabolism and adipogenesis during adipocyte maturation and showed that compound 1 substantially inhibited lipid accumulation compared to the control. Consistently, the expression of the adipocyte marker gene (Adipsin) was reduced upon incubation with 1. Further, we evaluated the effects of 1 on lipid metabolism by measuring the transcription of lipolytic and lipogenic genes. The expression of the lipolytic gene ATGL was significantly elevated upon exposure to 1 during adipogenesis, whereas the expression of lipogenic genes FASN and SREBP1 was significantly reduced upon treatment with 1. Thus, our findings provide experimental evidence that the steroid derived from Hydractinia-associated C. sphaerospermum SW67 is a potential therapeutic agent for obesity.

Guideline for Extended Release Oral Dosage Forms : Development, Evaluation, and Application of In Vitro/In Vivo Correlations (서방성 경구제형의 개발과 평가 및 생체내.외 상관성 연구를 위한 가이드라인)

  • Choi, Sun-Ok;Jeong, Sung-Hee;Um, So-Young;Jung, Seo-Jeong;Kim, Joo-Il;Kim, Ok-Hee
    • Journal of Pharmaceutical Investigation
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    • v.35 no.6
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    • pp.471-481
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    • 2005
  • In Korea, generic drug and bioequivalence test are the hot issues since a new medical system of separation of dispensary from medical practice was started in 2000. The KFDA(Korea FDA) had revised several times ${\ulcorner}Guidance\;for\;bioequivalence\;test{\lrcorner}$. In vitro dissolution test has been extensively used as a quality control tool for solid oral dosage forms. In an effort to minimize unnecessary human testing, in vitro/in vivo correlations (IVIVC) between in vitro dissolution and in vivo bioavailability are increasingly becoming an integral part on extended release drug product development. The recently published US guidance, ${\ulcorner}Extended\;release\;oral\;dosage\;forms\;:\;development,\;evaluation,\;and\;application\;of\;in\;vitro/in\;vivo\;correlations{\lrcorner}$ will be helpful for us to make our own guideline.