• Title/Summary/Keyword: Reverse phase HPLC

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Serum Concentrations of $\alpha$-Tocopherol, Carotenoids and Retinol of Normal Koreans

  • Lee, Yang-Cha-Kim;Kim, Mi-Kyung
    • Nutritional Sciences
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    • v.1 no.1
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    • pp.61-69
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    • 1998
  • Five hundred and seventy-eight healthy subjects (351 men and 227 women) with a mean age of 44.8 years (45.2 for men, 44.3 for women) participated in this study. The serum concentrations of $\alpha$-tocopherol, \beta$-carotene, $\alpha$-carotene, zeaxanthin + lutein, cryptoxanthin, lycopene and retinol of normal Koreans were measured, and their relation to gender, age, BMI, alcohol consumption, cigarette smoking, and menopausal status were evaluated. The concentrations of $\alpha$-tocopherol, carotenoids and retinol in serum were measured simultaneously by reverse phase HPLC (multi-wavelength, gradient and computerized automatic system). Average serum levels of $\alpha$-tocopherol, \beta$-carotene and retinol were 10.1$\pm$0.41\mu$g/dl, 33.1$\pm$1.24\mu$g/dl, and 82.0$\pm$1.63\mu$g/dl for men and 11.1$\pm$0.74\mu$g/ml, 48.1$\pm$1.60\mu$g/dl and 64.5$\pm$1.96\mu$g/dl for women, respectively, Serum concentrations of $\alpha$-tocopheol \beta$-carotene, $\alpha$-carotene, cryptoxanthin and lycopene appeared to be higher in women than in men. The serum concentrations of zeaxanthin + lutein increased with the increase in age of men and those of $\alpha$-carotene and \beta$-carotene increased with the increase in age of women. For men, current smokers showed significantly lower serum concentrations of \beta$-carotene and $\alpha$-carotene than ex-smokers (p <0.05) . Current drinkers also showed significantly lower serum \beta$-carotene and zeaxanthin + lutein concentrations than ex-drinkers. For women, current smokers showed significantly love. serum concentrations of zeaxanthin+lutein and cryptoxanthin than ex-smokes (p<0.05). Men with BMI$\geq$24 showed significantly lower serum concentrations of \beta$-carotene, $\alpha$-carotene, lycopene and cryptoxanthin than men with BMI < 20. The mean concentrations of $\alpha$-tocopherol, \beta$-carotene, $\alpha$-carotene and lycopene far postmenopausal women were higher than those for premenopausal women (p<0.05). In conclusion, there were obvious differences in serum $\alpha$-tocopherol and carotenoids levels depending on gender, age, cigarette-smoking habits, alcohol consumption, BMI and menopausal status. further studies are required to establish the normal levels of these vitamins for children and the elderly and to elucidate their roles in different disease states.

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Synthesis and Evaluation of F-18 Labeled 2'-Deoxy-2'-fluoro-5-methyl-1-β-L-arabinofuranosyluracil (L-[18F]FMAU)

  • Jo, Nam-Hyun;Moon, Byung-Seok;Hong, Su-Hee;An, Gwang-Il;Choi, Tae-Hyun;Cheon, Gi-Jeong;Cho, Jung-Hyuck;Yoo, Kyung-Ho;Lee, Kyo-Chul;Oh, Chang-Hyun
    • Bulletin of the Korean Chemical Society
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    • v.28 no.12
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    • pp.2449-2453
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    • 2007
  • L-[18F]FMAU ([18F]1b) was prepared from the precursor 2-O-[(trifluoromethyl)-sulfonyl]-1,3,5-tri-Obenzoyl- α-L-ribofuranose, by coupling the radioactive fluoro-sugar with the corresponding silylated thymine in 4 steps. The final products, including the α and β anomers, were purified using reverse phase HPLC with an appropriate solvent (5% CH3CN/H2O) at a flow rate of 3.0 mL/min. The total elapsed time of synthesis was about 180-200 min from EOB. The α/β anomeric ratio of the compounds was about 1:9, and the radiochemical purity of the product (β-form) was >98% with decay-corrected yields of 25-35%. All radioactive samples were confirmed using co-injection with pure non-radioactive analogues in every step. In the cellular uptake in vitro test of herpes simplex virus-thymidine kinase (HSV1-TK) gene expressed cells, the percent uptake of injected dose (%ID) of L- and D-FMAU was 37.28 and 65.86, respectively after 240 min incubation. However, the relative uptake (MCA-TK/MCA cellular uptake ratio) of L-FMAU was higher than that of D-FMAU (%ID of L-FMAU, 0.36 and D-FMAU, 0.93 after 240 min incubation in MCA cells). This means that L-FMAU will show better specific HSV1-TK gene expressed cell uptake for selective HSV1-TK gene imaging.

Biodistribution of 99mTc Labeled Integrin Antagonist

  • Jang, Beom-Su;Park, Seung-Hee;Shin, In Soo;Maeng, Jin-Soo;Paik, Chang H.
    • Toxicological Research
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    • v.29 no.1
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    • pp.21-25
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    • 2013
  • The selective targeting of an integrin ${\alpha}_v{\beta}_3$ receptor using radioligands may enable the assessment of angiogenesis and integrin ${\alpha}_v{\beta}_3$ receptor status in tumors. The aim of this research was to label a peptidomimetic integrin ${\alpha}_v{\beta}_3$ antagonist (PIA) with $^{99m}Tc(CO)_3$ and to test its receptor targeting properties in nude mice bearing receptor-positive tumors. PIA was reacted with tris-succinimidyl aminotriacetate (TSAT) (20 mM) as a PIA per TSAT. The product, PIA-aminodiacetic acid (ADA), was radiolabeled with $[^{99m}Tc(CO)_3(H_2O)_3]^{+1}$, and purified sequentially on a Sep-Pak C-18 cartridge followed by a Sep-Pak QMA anion exchange cartridge. Using gradient C-18 reverse-phase HPLC, the radiochemical purity of $^{99m}Tc(CO)_3$-ADA-PIA (retention time, 10.5 min) was confirmed to be > 95%. Biodistribution analysis was performed in nude mice (n = 5 per time point) bearing receptor-positive M21 human melanoma xenografts. The mice were administered $^{99m}Tc(CO)_3$-ADA-PIA intravenously. The animals were euthanized at 0.33, 1, and 2 hr after injection for the biodistribution study. A separate group of mice were also co-injected with 200 ${\mu}g$ of PIA and euthanized at 1 hr to quantify tumor uptake. $^{99m}Tc(CO)_3$-ADA-PIA was stable in phosphate buffer for 21 hr, but at 3 and 6 hr, 7.9 and 11.5% of the radioactivity was lost as histidine, respectively. In tumor bearing mice, $^{99m}Tc(CO)_3$-ADA-PIA accumulated rapidly in a receptor-positive tumor with a peak uptake at 20 min, and rapid clearance from blood occurring primarily through the hepatobiliary system. At 20 min, the tumor-to-blood ratio was 1.8. At 1 hr, the tumor uptake was 0.47% injected dose (ID)/g, but decreased to 0.12% ID/g when co-injected with an excess amount of PIA, indicating that accumulation was receptor mediated. These results demonstrate successful $^{99m}TC$ labeling of a peptidomimetic integrin antagonist that accumulated in a tumor via receptor-specific binding. However, tumor uptake was very low because of low blood concentrations that likely resulted from rapid uptake of the agent into the hepatobiliary system. This study suggests that for $^{99m}Tc(CO)_3$-ADA-PIA to be useful as a tumor detection agent, it will be necessary to improve receptor binding affinity and increase the hydrophilicity of the product to minimize rapid hepatobiliary uptake.

Purification of a Protease Produced by Bacillus subtilis PCA 20-3 Isolated from Korean Traditional Meju (전통 메주로부터 분리한 Bacillus subtilis PCA 20-3 유래 Protease 의 정제)

  • Lim, Seong-Il;Yoo, Jin-Young
    • Korean Journal of Food Science and Technology
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    • v.31 no.6
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    • pp.1635-1641
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    • 1999
  • Bacillus subtilis PCA20-3 was isolated from meju and was found to produce a protease. The strain produced the maximum amount of enzyme in the medium containing soytone (0.2%), soluble starch (2%), $(NH_4)_2SO_4\;(0.1%),\;CaCl_2(0.1%),\;yeast\;extract\;(0.01%),\;K_2HPO_4\;(0.1%),\;and\;KH_2PO_4\;(0.1%)$. Protease was first concentrated by ammonium sulfate (80% saturation, w/v) precipitation of culture supernatant. Then the enzyme was purified by column chromatography using CM Sephadex C-50. The collected proteins were rechromatographed using Sephadex G-100 gel filtration column. The fraction with protease active from Sephadex G-100 gel chromatography was found to be pure when examined by SDS-polyacrylamide gel electrophoresis and YMC-pak reverse phase chromatography. Specific activity, yield and purity were 76 U/mg. 2.7%, and 7.6 fold, respectively. The molecular weight of the enzyme was estimated to be 31.5 kDa by SDS-PAGE. The number of amino acids calculated from molecular weight was evaluated about 321 residues. N-terminal sequence of the enzyme was $Val^1-Pro^2-Tyr^3-Gly^4-Val^5-Ser^6-Gln^7-Gly^8-Lys^9-Ala^{10}$.

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Enzymatic Synthesis of Low Trans Fats Using Rice Bran Oil, Palm Stearin and High Oleic Sunflower Seed Oil (미강유, 팜스테아린 및 고올레인산 해바라기씨유를 이용한 저트랜스 유지의 효소적 합성)

  • Kim, Ji-Young;Lee, Ki-Teak
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.38 no.4
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    • pp.470-478
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    • 2009
  • Low trans fats were synthesized by interesterification of rice bran oil (RBO), palm stearin (PS) and high oleic sunflower seed oil (HO) using TLIM from Thermomyces lanuginosa. After 24-h reaction, physicochemical characteristics such as fatty acid and triacylglycerol composition, solid fat content, melting point, tocopherol, oryzanol and phytosterol contents were evaluated. Trans fatty acid contents of the produced low-trans fats showed less than 0.5 wt%. Mostly, triacylglycerol species in the products were palmitoyl-linoleoyl-oleoyl-glycerol (PLO), palmitoyl-oleoyl-oleoyl-glycerol (POO) and palmitoyl-oleoyl-palmitoyl-glycerol (POP). Total tocopherol contents ranged from 6.94 to 11.83 mg/100 g while $0.18{\sim}0.49$ mg/100 g of $\gamma$-oryzanol and $182.47{\sim}269.08$ mg/100 g of phytosterols were observed depending on the substrates ratios. When the content of PS in the reaction substrate was increased, solid fat content and slip melting points were increased.

Dammarane-type triterpene oligoglycosides from the leaves and stems of Panax notoginseng and their antiinflammatory activities

  • Li, Juan;Wang, Ru-Feng;Zhou, Yue;Hu, Hai-Jun;Yang, Ying-Bo;Yang, Li;Wang, Zheng-Tao
    • Journal of Ginseng Research
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    • v.43 no.3
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    • pp.377-384
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    • 2019
  • Background: Inflammation is widespread in the clinical pathology and closely associated to the progress of many diseases. Triterpenoid saponins as a key group of active ingredients in Panax notoginseng (Burk.) F.H. Chen were demonstrated to show antiinflammatory effects. However, the chemical structures of saponins in the leaves and stems of Panax notoginseng (PNLS) are still not fully clear. Herein, the isolation, purification and further evaluation of the antiinflammatory activity of dammarane-type triterpenoid saponins from PNLS were conducted. Methods: Silica gel and reversed-phase C8 column chromatography were used. Furthermore, preparative HPLC was used as a final purification technique to obtain minor saponins with high purities. MS, NMR experiments, and chemical methods were used in the structural identifications. The antiinflammatory activities of the isolated saponins were assessed by measuring the nitric oxide production in RAW 264.7 cells stimulated by lipopolysaccharides. Real-time reverse transcription polymerase chain reaction was used to measure the gene expressions of inflammation-related gene. Results: Eight new minor dammarane-type triterpene oligoglycosides, namely notoginsenosides LK1-LK8 (1-8) were obtained from PNLS, along with seven known ones. Among the isolated saponins, gypenoside IX significantly suppressed the nitric oxide production and inflammatory cytokines including tumor necrosis $factor-{\alpha}$, interleukin 10, interferon-inducible protein 10 and $interleukin-1{\beta}$. Conclusion: The eight saponins may enrich and expand the chemical library of saponins in Panax genus. Moreover, it is reported for the first time that gypenoside IX showed moderate antiinflammatory activity.

Antimicrobial effect of Enterococcus faecalis BMSE-HMP005 isolated from human breast milk against multidrug-resistant bacteria (모유 유래 유산균 Enterococcus faecalis BMSE-HMP005의 다제내성 균에 대한 항균효과)

  • Lee, Jeong-Eun;Kim, Soo-bin;Yu, Du-na;Jo, So-Yeon;Kim, Ae-Jung;Kook, Moochang
    • Korean Journal of Food Science and Technology
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    • v.54 no.2
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    • pp.209-217
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    • 2022
  • In this study, Enterococcus faecalis BMSE-HMP005 isolated from human breast milk demonstrated antimicrobial effects against multidrug-resistant (MDR) bacterial strains. The bacteriocin produced by E. faecalis BMSE-HMP005 was fractionated using reverse-phase high-performance liquid chromatography. This fraction showed antimicrobial effects against both gram-positive and gram-negative MDR bacteria. No hemolytic reactions were observed. E. faecalis BMSEHMP005 was resistant to vancomycin; however, kanamycin, ampicillin, and erythromycin showed minimum inhibitory concentrations that were lower than the acceptable range provided by the European Food Safety Authority. For artificial gastric juice and bile acid, the survival rates were 98.67% and 95.70%, respectively. These results show the potential utility of E. faecalis BMSE-HMP005 as a probiotic with remarkable antimicrobial effects against MDR bacteria.

Development and Validation of the Determination of Sorafenib in Human Plasma using Tandem Mass Spectrometry Coupled with Liquid Chromatography (고속액체크로마토그래피 텐덤질량분석기법을 이용한 사람 혈장 내 소라페닙 농도분석법의 개발 및 검정)

  • Park, Daejin;Lee, Sunggon;Kim, Woomi
    • Journal of Life Science
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    • v.22 no.11
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    • pp.1456-1462
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    • 2012
  • Sorafenib is a multikinase inhibitor and an oral anticancer drug approved for the treatment of patients with advanced renal cell carcinoma and those with unresectable hepatocellular carcinoma. The purpose of this study was to develop an efficient method of the determination of sorafenib in human plasma using tandem mass spectrometry coupled with liquid chromatography (LC/MS/MS) and validate the method by the guidelines of the Korean Food and Drug Administration (KFDA). Plasma samples ($100{\mu}l$) were added with chlorantraniliprole as an internal standard and then mixed with the 0.1% formic acid-containing extraction solution composed of isopropyl alcohol and ethyl acetate (1:4, v/v). After centrifugation, the supernatant was concentrated at $45^{\circ}C$ under negative pressure and centrifugal force. The residue was reconstituted with a mobile phase and injected into the HPLC instrument using a reverse phase Waters XTerra$^{TM}$ C18 column (particle size $3.5{\mu}m$). Liquid chromatography was carried out within the run time of 5 min using a mobile phase composed of buffer (0.1% formic acid and 10 mM ammonium formate), methanol, and acetonitrile (1:6:3, v/v/v). The analytes were monitored by tandem mass spectrometry in the multiple reaction monitoring method programmed to detect sorafenib at 'm/z 465.2 ${\rightarrow}$ 252.5' and chlorantraniliprole at 'm/z 484.4 ${\rightarrow}$ 286.2' with positive electrospray ionization mode ($ES^+$). The result showed the proper linearity ($r^2$ > 0.99) over the range of 2,000-5,000 ng/ml with good accuracy (90.7-103.9%) and precision (less than 10%). The newly developed method using LC/MS/MS was validated by the guideline of KFDA and identified as more sensitive compared to the previous methods.

Studies on Simultaneous Determination of Chlorophyll a and b, Pheophorbide a, and $\beta-Carotene$ in Chlorella and Spirulina Products (클로렐라 및 스피루리나제품에 함유된 엽록소 a, b, 페오포르바이드 a 및 $\beta$-카로틴의 동시분석법에 관한 연구)

  • Lee Young Ja;Kim So Hee;Kim Jin-Sook;Han Jeong A;Seo Hae Jeom;Lim Hyo Jeong;Choi Soo Young
    • Journal of Food Hygiene and Safety
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    • v.20 no.3
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    • pp.141-146
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    • 2005
  • A simple and sensitive analysis method based on reverse phase (RP) HPLC with UV detector was developed for simultaneous determination of chlorophyll a and b, pheophorbide a and $\beta-Carotene$ in Chlorella and Spirulina products. For added concentration $(50\;\mug/ml)$ of chlorophyll a and b, pheophorbide a and $\beta-Carotene$, recoveries of those were 70.3, 71.6, 60.1 and $90.5\%$, respectively, with relative standard deviations of 2.8,6.0, 10.6 and $10.4\%$. Limit of detection and quantification had ranges of $0.1\sim1.0\;\mug/ml$ and $0.2\sim2.0\;\mug/ml$, respectively. Calibration curve was linear with correlation coefficient of 0.995 for chlorophyll a and b, pheophorbide a and $\beta-Carotene$. Results of simultaneous determination in Chlorella and Spirulina products were showed ranges of $121.g\sim543.0\;\mug/ml$ for chlorophyll a,$0.6\sim160.0\;\mug/ml$ for chlorophyll b, $19.2\sim60.3\;\mug/ml$ for pheophorbide a and $383.6\sim1713.7\;\mug/ml$ for $\beta-Carotene$, respectively. Chlorophyll b contents in Chlorella products were detected above 30 times level to those in Spirulina products. $\beta-Carotene$ contents in Spirulina products were detected 2.7 times level to those in Chlorella products.

Changes in Polyamine and Tyramine Concentrations in Rice (Oryza sativa L.) during Maturation and Preharvest Sprouting (벼 등숙기와 수발아 기간동안 폴리아민과 티라민의 농도변화)

  • Kim, Tae-Wan;Kim, Jae-Hun;Hwang, Seon-Woong;Hong, Byuong-Hee;Lee, Sang-Eun;Yun, Seung-Gil;An, Jae-Ho
    • Korean Journal of Soil Science and Fertilizer
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    • v.38 no.1
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    • pp.1-7
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    • 2005
  • The levels of polyamines were measured to investigate the alternative nitrogen metabolism during maturation and sprouting in rice. The rice plants (cv. Ansanbyeo) were cultivated in 20-year-old non-fertilized field. The flag leaves and spikes were collected weekly after the earing stage and the seeds were harvested daily after lodging. Free, bound, and conjugated polyamines were analyzed using reverse phase HPLC. Putrescine, spermidine, spermine, agmatine and tyramine were the major amines found in rice. The level of stress-induced amine, putrescine increased during the preharvest sprouting confirming that the process was a stress to the plants. With all other polyamines, tyramine in free form decreased in flag leaves and panicles during seed maturation. However, agmatine in bound form showed a noticeable increase about 8-fold during 6 weeks period of maturation after which it declined to the bottom level. Among the individual amines, tyramine and spermine in conjugated form showed a marked change during matutation and sprouting. Interestingly, the level of tyramine with all conjugated polyamine decreased in spikes during seed maturation and increased during preharvest sprouting implying that tyrosine decarboxyation and conjugation to phenolic acids may play a key role in preharvest sprouting. Spermine in conjugated form was synthesized only at the early earing stage in the level of $3.4mole\;g^{-1}$ fresh weight, and then decreased to the level of nmole during maturation. Thereafter, it dramatically increased to 2.8 mole during preharvest sprouting. In this study we found the tyramine is a major amine in rice, and it would play a critical role in N-assimilation during seed maturation and sprouting.