• Title/Summary/Keyword: high performance liquid chromatography

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Determination of Thiamine in Pharmaceutical Preparations by Reverse Phase Liquid Chromatography Without Use of Organic Solvent

  • Suh, Joon Hyuk;Kim, Junghyun;Jung, Juhee;Kim, Kyunghyun;Lee, Seul Gi;Cho, Hyun-Deok;Jung, Yura;Han, Sang Beom
    • Bulletin of the Korean Chemical Society
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    • v.34 no.6
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    • pp.1745-1750
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    • 2013
  • A novel green aqueous mobile phase modified with room temperature ionic liquids (RTILs) was employed in the absence of volatile organic solvents or ion-pairing reagents to analyze thiamine, a very polar compound, by reverse phase high performance liquid chromatography (RP-HPLC). Due to its strongly hydrophilic nature, thiamine was eluted near the column dead time ($t_0$) using a mobile phase without adding RTILs or ion-pairing reagents, even if a 100% aqueous mobile phase, which has weak elution power under reverse phase conditions, was used. Thus, 1-ethyl-3-methyl-imidazolium hexafluorophosphate ([EMIM][$PF_6$]), which has the strongest chaotropic effect, was selected as a mobile phase additive to improve retention and avoid baseline disturbances at $t_0$. Various mobile phase parameters such as cation moiety, chaotropic anion moiety, pH and concentration of RTILs were optimized to determine thiamine at the proper retention time. Method validation was performed to assess linearity, intra- and inter-day accuracy and precision, recovery and repeatability; all results were found to be satisfactory. The developed method was also compared to the current official United States Pharmacopoeia (USP) and Korean Pharmacopoeia (KP) methods using an organic mobile phase containing an ionpairing reagent by means of evaluating various chromatographic parameters such as the capacity factor, theoretical plate number, peak asymmetry and tailing factor. The results indicated that the proposed method exhibited better efficiency of thiamine analysis than the official methods, and it was successfully applied to quantify thiamine in pharmaceutical preparations.

Analysis of oligosaccharides from Panax ginseng by using solid-phase permethylation method combined with ultra-high-performance liquid chromatography-Q-Orbitrap/mass spectrometry

  • Li, Lele;Ma, Li;Guo, Yunlong;Liu, Wenlong;Wang, Yang;Liu, Shuying
    • Journal of Ginseng Research
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    • v.44 no.6
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    • pp.775-783
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    • 2020
  • Background: The reports about valuable oligosaccharides in ginseng are quite limited. There is an urgent need to develop a practical procedure to detect and analyze ginseng oligosaccharides. Methods: The oligosaccharide extracts from ginseng were permethylated by solid-phase methylation method and then were analyzed by ultra-high-performance liquid chromatography-Q-Orbitrap/MS. The sequence, linkage, and configuration information of oligosaccharides were determined by using accurate m/z value and tandem mass information. Several standard references were used to further confirm the identification. The oligosaccharide composition in white ginseng and red ginseng was compared using a multivariate statistical analysis method. Results: The nonreducing oligosaccharide erlose among 12 oligosaccharides identified was reported for the first time in ginseng. In the comparison of the oligosaccharide extracts from white ginseng and red ginseng, a clear separation was observed in the partial least squares-discriminate analysis score plot, indicating the sugar differences in these two kinds of ginseng samples. The glycans with variable importance in the projection value large than 1.0 were considered to contribute most to the classification. The contents of oligosaccharides in red ginseng were lower than those in white ginseng, and the contents of maltose, maltotriose, maltotetraose, maltopentaose, maltohexaose, maltoheptaose, maltooctaose, maltononaose, sucrose, and erlose decreased significantly (p < 0.05) in red ginseng. Conclusion: A solid-phase methylation method combined with liquid chromatography-tandem mass spectrometry was successfully applied to analyze the oligosaccharides in ginseng extracts, which provides the possibility for holistic evaluation of ginseng oligosaccharides. The comparison of oligosaccharide composition of white ginseng and red ginseng could help understand the differences in pharmacological activities between these two kinds of ginseng samples from the perspective of glycans.

Qualitative Analysis of the Major Constituents in Traditional Oriental Prescription Bang-poong-tong-sung-san by Liquid Chromatography/Ultraviolet Detector/Ion-Trap Time-of-Flight Mass Spectrometry

  • Eom, Han Young;Kim, Hyung-Seung;Han, Sang Beom
    • Mass Spectrometry Letters
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    • v.5 no.1
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    • pp.24-29
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    • 2014
  • An advanced and reliable high performance liquid chromatography (HPLC)/ultraviolet detector (UV)/ion-trap time-of-flight (IT-TOF) mass spectrometry was developed for the simultaneous quantification of 19 marker compounds in Bang-poong-tong-sung-san (BPTS), a traditional oriental prescription. Various parameters affecting HPLC separation and IT-TOF detection were investigated, and optimized conditions were identified. The separation was achieved on a Capcell PAK C18 column ($1.5mm{\times}250mm$, $5{\mu}m$ particle size) using a gradient elution of acetonitrile and water containing 0.1% formic acid at a flow rate of 0.1 mL/min. The column temperature was maintained at $40^{\circ}C$ and the injection volume was $2{\mu}L$. IT-TOF system was equipped with an electrospray ion source (ESI) operating in positive or negative ion mode. The optimized electrospray ionization parameters were as follows: ion spray voltage, +4.5 kV (positive ion mode), or -3.5 kV (negative ion mode); drying gas ($N_2$), 1.5 L/min; heat block temperature, $200^{\circ}C$. Automatic $MS^n$ (n = 1~3) analyses were carried out to obtain structural information of analytes. Elemental compositions and their mass errors were calculated based on their accurate masses obtained from a formula predictor software. The marker compounds in BPTS were identified by comparisons between $MS^n$ spectra from standards and those from extracts. Moreover, the libraries of $MS^2$ and $MS^3$ spectra and accurate masses of parent and fragment ions for marker compounds were constructed. The developed method was successfully applied to the BPTS extracts and identified 17 out of 19 marker compounds in the BPTS extracts.

The analysis of the urushiol congeners from the extracts of lacquer trees (옻나무 추출물 중 우루시올 동종체의 함량비 분석)

  • Cho, Yumi;Jung, Yu-Kyung;Kim, Jinsan;Lee, Joonbae;Paeng, Ki-Jung
    • Analytical Science and Technology
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    • v.22 no.1
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    • pp.65-74
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    • 2009
  • Active components of lacquer tree referred to as urushiol congeners, which are catechol derivatives with various alkyl or alkenyl substituents. The olefin side chains typically have one, two or three double bonds. In this study, the each congener's ratio analysis of extracts from korean lacquer tree are compared to the one from other asian lacquer tree. Extraction was performed using liquid-liquid extraction (LLE) method with soxhlet system from tree's bark and sap. Extracts were analyzed by reverse phase liquid chromatography and on-line electro spray ionization mass spectrometry (LC-MS/MS).

Purification and Characterization of Cop, a Protein Involved in the Copy Number Control of Plasmid pE194

  • Kwak, Jin-Hwan;Kim, Jung-Ho;Kim, Mu-Yong;Choi, Eung-Chil
    • Archives of Pharmacal Research
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    • v.21 no.3
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    • pp.291-297
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    • 1998
  • Cop protein has been overexpressed in Escherichia coli using a T7 RNA polymerase system. Purification to apparent homogeneity was achieved by the sequential chromatography on ion exchange, affinity chromatography, and reverse phase high performance liquid chromatography system. The molecular weight of the purified Cop was estimated as 6.1 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). But the molecular mass of the native state Cop was shown to be 19 kDa by an analytical high performance size exclusion chromatography, suggesting a trimer-like structure in 50 mM Tris-HCI buffer (pH 7.5) containing 100 mM NaCl. Cop protein Was calculated to contain $39.1% {\alpha}-helix, 16.8% {\beta}-sheet$, 17.4% turn, and 26.8% random structure. The DNA binding property of Cop protein expressed in E. coli Was preserved during the expression and purification process. The isoelectric point of Cop was determined to be 9.0. The results of amino acid composition analysis and N-terminal amino acid sequencing of Cop showed that it has the same amino acid composition and N-terminal amino acid sequence as those deduced from its DNA sequence analysis, except for the partial removal of N-terminal methionine residue by methionyl-aminopeptidase in E. coli.

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Determination of Ginsenosides Content in Korean Ginseng Seeds and Roots by High Performance Liquid Chromatography

  • Hu, Jiang Ning;Lee, Jeung-Hee;Shin, Jung-Ah;Choi, Jae-Eul;Lee, Ki-Teak
    • Food Science and Biotechnology
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    • v.17 no.2
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    • pp.430-433
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    • 2008
  • A high performance liquid chromatography (HPLC) method has been successfully developed to identify and quantify major ginsenosides in Korean ginseng seeds and roots. Using gradient elution of acetonitrile and water without buffer, the 6 major ginsenosides ($Rb_1,\;Rb_2$, Rc, Rd, Re, and $Rg_1$) were identified. Compared with ginseng roots, the amount of ginsenoside Re and Rd in ginseng seeds were significantly higher than those in ginseng roots (p<0.05). In ginseng seeds, the content of protopanaxtriol (PPT) was higher than that of protopanaxdiol (PPD) and the ratio of PPT and PPD was approximately 2.2 : 1. However, the content of PPT was lower than that of PPD in ginseng roots. It should be mentioned that both content of PPT and PPD in ginseng seeds were much higher than those in ginseng roots.

Studies on Rooting of Taxus cuspidata Cuttings by Electric Treatment (전기처리에 의한 주목의 삽목발근에 관한 연구)

  • 정진철;최정호;장규관
    • Journal of the Korean Institute of Landscape Architecture
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    • v.22 no.2
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    • pp.123-132
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    • 1994
  • Tow-year-old Taxus cuspidata shoot cuttings were treated with various electrical impacts of cathode on their base and anode on their apex by normal and reverse source. The cuttings were previously treated with 200ppm IBA for 12 hours and the rooting percentage, the length, and the weight of roots were checked. The auxin contents of cuttings were also examined by high-performance liquid chromatography. The results obtained on this study are as follows; 1. Normal treatment, cathode into the base and anode into apex, seemed to accelerate rooting while reverse treatment showed less effective than normal treatment on rooting, but both treatments were more effective than control. 2. The impact of electrical treatment at 30mV for 30min has a remarkable effect on the percentage rooting, the length, and the weight of roots. 3. Root primordia were formed at the basipetal end of cuttings where the end of primary pith ray meets the cambium in control treatment and formed at the basal part of cuttings irregularly in electrical treatment. 4. High-performance liquid chromatography showed electrical treatment was more effective on auxin accumulation than control, and 30mV-30min was the most effective on auxin accumulation.

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