• Title/Summary/Keyword: LC/MS analysis

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Lessons from the Sea : Genome Sequence of an Algicidal Marine Bacterium Hahella chehuensis (적조 살상 해양 미생물 Hahella chejuensis의 유전체 구조)

  • Jeong Hae-Young;Yoon Sung-Ho;Lee Hong-Kum;Oh Tae-Kwang;Kim Ji-Hyun
    • Microbiology and Biotechnology Letters
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    • v.34 no.1
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    • pp.1-6
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    • 2006
  • Harmful algal blooms (HABs or red tides), caused by uncontrolled proliferation of marine phytoplankton, impose a severe environmental problem and occasionally threaten even public health. We sequenced the genome of an EPS-producing marine bacterium Hahella chejuensis that produces a red pigment with the lytic activity against red-tide dinoflagellates at parts per billion level. H. chejuensis is the first sequenced species among algicidal bacteria as well as in the order Oceanospirillales. Sequence analysis indicated a distant relationship to the Pseudomonas group. Its 7.2-megabase genome encodes basic metabolic functions and a large number of proteins involved in regulation or transport. One of the prominent features of the H. chejuensis genome is a multitude of genes of functional equivalence or of possible foreign origin. A significant proportion (${\sim}23%$) of the genome appears to be of foreign origin, i.e. genomic islands, which encode genes for biosynthesis of exopolysaccharides, toxins, polyketides or non-ribosomal peptides, iron utilization, motility, type III protein secretion and pigment production. Molecular structure of the algicidal pigment was determined to be prodigiosin by LC-ESI-MS/MS and NMR analyses. The genomics-based research on H. chejuensis opens a new possibility for controlling algal blooms by exploiting biotic interactions in the natural environment and provides a model in marine bioprospecting through genome research.

Simultaneous qualitative and quantitative analysis of morroniside and hederacoside D in extract mixture of Cornus officinalis and Stauntonia hexaphylla leaves to improve benign prostatic hyperplasia by HPLC-UV

  • Dan, Gao;Cho, Chong Woon;Vinh, Le Ba;Kim, Jin Hyeok;Cho, Kyoung Won;Kim, Young Ho;Kang, Jong Seong
    • Analytical Science and Technology
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    • v.33 no.5
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    • pp.224-231
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    • 2020
  • With the improvement in the standard of living and extension of life expectancy, the incidence of prostate diseases has increased yearly, thus becoming a serious disease affecting the health of men. The extract mixture of Cornus officinalis and Stauntonia hexaphylla leaves is a developed functional food formula to improve prostate health. This study developed a simultaneous analytical method of bioactive compounds for quantifying the mixture of Cornus officinalis and S. hexaphylla leaves using high-pressure liquid chromatography-ultraviolet (HPLC-UV). HPLC analytical condition was performed on a Hector C18 column with a mobile phase of 0.1 % formic acid in water (A) and 0.1 % formic acid in acetonitrile (B) under the following gradient conditions: 0-50 min, 12 %-40 % (B) at a flow rate of 1.0 mL/min. Meanwhile, this method was validated properly and successfully used to quantify the bioactive components of morroniside and hederacoside D in 20 sample batches and assess the quality of different ages and seasons of S. hexaphylla leaves. The result showed that the content of morroniside in the extract mixture of Cornus officinalis and S. hexaphylla leaves ranged from 1.38-1.62 mg/g, and the hederacoside D ranged from 28.42-32.02 mg/g, suggesting that this novel analytical method will be suitable for the quality control of the extract mixture to improve benign prostatic hyperplasia.

Characterization of Xylanase from Lentinus edodes M290 Cultured on Waste Mushroom Logs

  • Lee, Jae-Won;Gwak, Ki-Seob;Kim, Su-Il;Kim, Mi-Hyang;Choi, Don-Ha;Choi, In-Gyu
    • Journal of Microbiology and Biotechnology
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    • v.17 no.11
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    • pp.1811-1817
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    • 2007
  • Extracellular enzymes from Lentinus edodes M290 on normal woods (Quercus mongolica) and waste logs from oak mushroom production were comparatively investigated. Endoglucanase, cellobiohydrolase, ${\beta}$-glucosidase, and xylanase activities were higher on waste mushroom logs than on normal woods after 1. edodes M290 inoculation. Xylanase activity was especially different, with a three times higher activity on waste mushroom logs. When the waste mushroom logs were used as a carbon source, a new 35 kDa protein appeared. After the purification, the optimal pH and temperature for xylanase activity were determined to be 4.0 and $50^{\circ}C$, respectively. More than 50% of the optimal xylanase activity was retained when the temperature was increased from 20 to $60^{\circ}C$, after a 240 min reaction. At $40^{\circ}C$, the xylanase maintained 93% of the optimal activity, after a 240 min reaction. The purified xylanase showed a very high homology to the xylanase family 10 from Aspergillus terreus by LC/MS-MS analysis. The highest Xcorr (1.737) was obtained from the peptide KWI SQGIPIDGIG SQTHLGSGGS WTVK originated from Aspergillus terreus, indicating that the 35 kDa protein was xylanase. This protein showed low homology to a previously reported L. edodes xylanase sequence.

Comparative Proteomic Profiling of Pancreatic Ductal Adenocarcinoma Cell Lines

  • Kim, Yikwon;Han, Dohyun;Min, Hophil;Jin, Jonghwa;Yi, Eugene C.;Kim, Youngsoo
    • Molecules and Cells
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    • v.37 no.12
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    • pp.888-898
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    • 2014
  • Pancreatic cancer is one of the most fatal cancers and is associated with limited diagnostic and therapeutic modalities. Currently, gemcitabine is the only effective drug and represents the preferred first-line treatment for chemotherapy. However, a high level of intrinsic or acquired resistance of pancreatic cancer to gemcitabine can contribute to the failure of gemcitabine treatment. To investigate the underlying molecular mechanisms for gemcitabine resistance in pancreatic cancer, we performed label-free quantification of protein expression in intrinsic gemcitabine-resistant and -sensitive human pancreatic adenocarcinoma cell lines using our improved proteomic strategy, combined with filter-aided sample preparation, single-shot liquid chromatography-mass spectrometry, enhanced spectral counting, and a statistical method based on a power law global error model. We identified 1931 proteins and quantified 787 differentially expressed proteins in the BxPC3, PANC-1, and HPDE cell lines. Bioinformatics analysis identified 15 epithelial to mesenchymal transition (EMT) markers and 13 EMT-related proteins that were closely associated with drug resistance were differentially expressed. Interestingly, 8 of these proteins were involved in glutathione and cysteine/methionine metabolism. These results suggest that proteins related to the EMT and glutathione metabolism play important roles in the development of intrinsic gemcitabine resistance by pancreatic cancer cell lines.

Effect of Gongjindon a Traditional Korean Polyherbal Formula, on the Pharmacokinetics Profiles of Donepezil in Male SDRats (1) -Single Oral Combination Treatment of Donepezil 10mg/kg with Gongjindan 100mg/kg within 5 min-

  • Chung, Dae-Kyoo;Kwon, Oh-Dae;Park, Soo-Jin;Lee, Young-Joon;Ku, Sae-Kwang
    • Journal of Society of Preventive Korean Medicine
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    • v.17 no.1
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    • pp.77-88
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    • 2013
  • Objectives : This study was aim to evaluate effects of pharmacodynamics and toxicity in combination therapy of donepezil with Gongjindan. Methods : After 10mg/kg of donepezil treatment, Gongjindan 100mg/kg was administered within 5 min. The plasma were collected at 30min before administration, 30min, 1, 2, 3, 4, 6, 8 and 24hrs after end of Gongjindan treatment, and plasma concentrations of donepezil were analyzed using LC-MS/MS methods. PK parameters of donepezil were analysis as compared with donepezil single administered rats. Results : Gongjindan markedly inhibited the absorption of donepezil regardless of sample time, from 30min to 8hrs after end of co-administration comparing with donepezil single treated rats. Especially the absorption of donepezil was significantly decreased at 2hrs after co-administration as compared with donepezil single treated rats, in the present study. Accordingly, the Cmax(-27.76%), $AUC_{0-t}$(-27.22%) and $AUC_{0-inf}$(-26.54%) of donepezil in co-administered rats were significantly decreased as compared with donepezil single treated rats, respectively. Conclusions : Based on the results of the present study, co-administration of Gongjindan decreases the oral bioavailability of donepezil by inhibiting the absorption. It is considered that the more detail pharmacokinetic studies should betested to conclude the effects of Gongjindan on the pharmacokinetics of donepezil, when they were co-administered, like the effects after co-administration with reasonable intervals considering the Tmax of donepezil and after repeated co-administrations.

Inhibition of Sphingolipid Metabolism Enhances Resveratrol Chemotherapy in Human Gastric Cancer Cells

  • Shin, Kyong-Oh;Park, Nam-Young;Seo, Cho-Hee;Hong, Seon-Pyo;Oh, Ki-Wan;Hong, Jin-Tae;Han, Sang-Kil;Lee, Yong-Moon
    • Biomolecules & Therapeutics
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    • v.20 no.5
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    • pp.470-476
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    • 2012
  • Resveratrol, a chemopreventive agent, is rapidly metabolized in the intestine and liver via glucuronidation. Thus, the pharmacokinetics of resveratrol limits its efficacy. To improve efficacy, the activity of resveratrol was investigated in the context of sphingolipid metabolism in human gastric cancer cells. Diverse sphingolipid metabolites, including dihydroceramides (DHCer), were tested for their ability to induce resveratrol cytotoxicity. Exposure to resveratrol ($100{\mu}M$) for 24 hr induced cell death and cell cycle arrest in gastric cancer cells. Exposure to the combination of resveratrol and dimethylsphingosine (DMS) increased cytotoxicity, demonstrating that sphingolipid metabolites intensify resveratrol activity. Specifically, DHCer accumulated in a resveratrol concentration-dependent manner in SNU-1 and HT-29 cells, but not in SNU-668 cells. LC-MS/MS analysis showed that specific DHCer species containing C24:0, C16:0, C24:1, and C22:0 fatty acids chain were increased by up to 30-fold by resveratrol, indicating that resveratrol may partially inhibit DHCer desaturase. Indeed, resveratrol mildly inhibited DHCer desaturase activity compared to the specific inhibitor GT-11 or to retinamide (4-HPR); however, in SNU-1 cells resveratrol alone exhibited a typical cell cycle arrest pattern, which GT-11 did not alter, indicating that inhibition of DHCer desaturase is not essential to the cytotoxicity induced by the combination of resveratrol and sphingolipid metabolites. Resveratrol-induced p53 expression strongly correlated with the enhancement of cytotoxicity observed upon combination of resveratrol with DMS or 4-HPR. Taken together, these results show that DHCer accumulation is a novel lipid biomarker of resveratrol-induced cytotoxicity in human gastric cancer cells.

Acrylamide Monitoring of Domestic Food Products (국내생산식품의 아크릴아마이드 함량분석)

  • Park, Jae-Young;Kim, Cheong-Tae;Kim, Hye-Young;Keum, Eun-Hee;Lee, Mi-Seon;Chung, So-Young;Sho, You-Sub;Lee, Jong-Ok;Oh, Sang-Suk
    • Korean Journal of Food Science and Technology
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    • v.36 no.6
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    • pp.872-878
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    • 2004
  • Food products selected based on their annual sales and international acrylamide research data were analyzed for quantitation of acrylamide. Samples including raw food, substitute meal, snack, drink, and sauce products were analyzed by LC/MS/MS methods adopted by PDA. Upon comparison, concentrations of acrylamide in these products were similar to those analyzed in other countries.

Residues Analysis of Acetamiprid, Boscalid, Imidacloprid and Pyraclostrobin in the Minor Crop Mustard Green under Greenhouse Conditions for Evaluation of their Potentiality of PLS Violation

  • Kim, Young Eun;Kim, Seon Wook;Lim, Da Jung;Kim, In Seon
    • Korean Journal of Environmental Agriculture
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    • v.39 no.3
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    • pp.214-221
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    • 2020
  • BACKGROUND: The demand for pesticide registration has kept increasing for minor crop cultivation in greenhouse since Positive List System (PLS) has been launched. Thus, much study on the evaluation of pesticide residues in minor crops is required to examine the demand. In this study, we evaluated residues of acetamiprid, boscalid, imidacloprid and pyraclostrobin in the minor crop mustard green to provide the potential data for their registration. METHODS AND RESULTS: Pesticide granule formulations of acetamiprid, boscalid, imidacloprid and pyraclostrobin were incorporated into soil and applied onto field soil surface at rates of 3 kg/10a, 6 kg/10a, 3 kg/10a and 6 kg/10a, respectively. The pesticides were also applied at the two times higher than the rates to compare the residues between the application rates. Mustard green seeds were sown 1 day after pesticide application and cultivated under greenhouse conditions. LC/MS/MS analyses coupled with a modified QuEChERs method were employed for determination of the pesticides in plant samples. The method limits of quantitation (LOQ) of the pesticides were 0.01 mg/kg, and the matrix calibration curves of the pesticides showed linearity with coefficient values of determination (r2) greater than 0.995. The average recovery values of the pesticides fortified in control samples at rates of LOQ and 10LOQ ranged from approximately 77.5% to 101.2% with relative standard deviation values lower than 14%. The pesticides in the mustard green samples cultivated for 53 days after sown were determined to be lower than the LOQ level. CONCLUSION: Acetamiprid, boscalid, imidacloprid and pyraclostrobin were found at a level lower than 0.01 mg/kg in the minor crop mustard green. Thus, their residues in mustard green would not violate PLS under greenhouse conditions.

Cloning, Purification, and Characterization of Recombinant Human Extracellular Superoxide Dismutase in SF9 Insect Cells

  • Shrestha, Pravesh;Yun, Ji-Hye;Kim, Woo Taek;Kim, Tae-Yoon;Lee, Weontae
    • Molecules and Cells
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    • v.39 no.3
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    • pp.242-249
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    • 2016
  • A balance between production and degradation of reactive oxygen species (ROS) is critical for maintaining cellular homeostasis. Increased levels of ROS during oxidative stress are associated with disease conditions. Antioxidant enzymes, such as extracellular superoxide dismutase (EC-SOD), in the extracellular matrix (ECM) neutralize the toxicity of superoxide. Recent studies have emphasized the importance of EC-SOD in protecting the brain, lungs, and other tissues from oxidative stress. Therefore, EC-SOD would be an excellent therapeutic drug for treatment of diseases caused by oxidative stress. We cloned both the full length (residues 1-240) and truncated (residues 19-240) forms of human EC-SOD (hEC-SOD) into the donor plasmid pFastBacHTb. After transposition, the bacmid was transfected into the Sf9-baculovirus expression system and the expressed hEC-SOD purified using FLAG-tag. Western blot analysis revealed that hEC-SOD is present both as a monomer (33 kDa) and a dimer (66 kDa), as detected by the FLAG antibody. A water-soluble tetrazolium (WST-1) assay showed that both full length and truncated hEC-SOD proteins were enzymatically active. We showed that a potent superoxide dismutase inhibitor, diethyldithiocarbamate (DDC), inhibits hEC-SOD activity.

Analysis of Iridoid Glycoside and GABA Content in the Roots of the Rehmannia glutinosa Cultivars (지황 품종별 뿌리에서 Iridoid 배당체와 GABA 분석)

  • Lee, Sang Hoon;Yoon, Jeong Su;Kim, Jae Kwang;Park, Chun Geun;Chang, Jae Ki;Kim, Yeon Bok
    • Korean Journal of Medicinal Crop Science
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    • v.25 no.3
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    • pp.146-151
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    • 2017
  • Background: Rehmannia glutinosa is a perennial herb belonging to the family Scrophulariaceae. Its roots have been utilized as a traditional medicine. The aim of this study was to elucidate the basic information of the roots of the R. glutinosa cultivars and their utilization. Methods and Results: The roots of R. glutinosa cultivars were harvested in the end of March. The two iridoid glycosides, aucubin and catalpol, were analyzed by liquid chromatography/mass spectrometry (LC/MS), whereas ${\gamma}-aminobutyric$ acid (GABA) was analyzed by gas chromatography/mass spectrometry (GC/MS). The aucubin content was the highest in the Dakang cultivar, whereas no aucubin was detected in the five cultivars. All cultivars had more than 12 mg/g catalpol content, and the maximum catalpol content was found in Jihwang 1. The GABA content was the highest in Suwon 1, and it was 40 times more than that in the Yeongang cultivar. Conclusions: The highest aucubin, catapol and GABA contents were detected in the Dakang, Jihwang 1, and Suwon 1, cultivars respectively. This study provides the crucial information regarding the versatile utilization and pedigree selection of R. glutinosa cultivars.