• Title/Summary/Keyword: HPLC(high performance liquid chromatography)

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Column cleaning, regeneration and storage of silica-based columns (실리카 기반 컬럼의 세척, 재생 및 보관 가이드)

  • Matt James;Mark Fever
    • FOCUS: LIFE SCIENCE
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    • no.1
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    • pp.1.1-1.4
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    • 2024
  • This article provides comprehensive guidance on the maintenance, cleaning, regeneration, and storage of silica-based HPLC (High-Performance Liquid Chromatography) columns. The general considerations emphasize the importance of using in-line filters and guard cartridges to protect columns from blockage and irreversible sample adsorption. While these measures help, contamination by strongly adsorbed sample components can still occur over time, leading to an increase in back pressure, loss of efficiency, and other issues. To maximize column lifetime, especially with UHPLC (Ultra-High Performance Liquid Chromatography) columns, it is advisable to use ultra-pure solvents, freshly prepared aqueous mobile phases, and to filter all samples, standards, and mobile phases. Additionally, an in-line filter system and sample clean-up on dirty samples are recommended. However, in cases of irreversible compound adsorption or column voiding, regeneration may not be possible. The document also provides specific recommendations for column cleaning procedures, including the flushing procedures for various types of columns such as reversed phase, unbonded silica, bonded normal phase, anion exchange, cation exchange, and size exclusion columns for proteins. The flushing procedures involve using specific solvents in a series to clean and regenerate the columns. It is emphasized that the flow rate during flushing should not exceed the specified limit for the particular column, and the last solvent used should be compatible with the mobile phase. Furthermore, the article outlines the storage conditions for silica based HPLC columns, highlighting the impact of storage conditions on the column's lifetime. It is recommended to flush all buffers, salts, and ion-pairing reagents from the column before storage. The storage solvent should ideally match the one used in the initial column test chromatogram provided by the manufacturer, and column end plugs should be fitted to prevent solvent evaporation and drying out of the packing bed.

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Simultaneous Gas Chromatographic Analysis of Ethanol and Acetic Acid in Vinegar (식초의 에탄올 및 초산 함량의 동시분석)

  • Yoon, Hee-Nam
    • Korean Journal of Food Science and Technology
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    • v.30 no.6
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    • pp.1247-1251
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    • 1998
  • A method is developed for simultaneously determining ethanol and acetic acid in vinegars by quantitative packed-column gas chromatography. Vinegars were filtrated and directly chromatographed on a $2\;m{\times}2\;mm$ stainless steel column packed with Tenax-GC, 80/100. Ethanol, isopropy alcohol as an internal standard, and acetic acid were completely separated within 20 min of running time without any interfering peaks. The accuracy of packed column gas solid chromatography (PCGSC) was discussed compared to the analytical data by titration, high performance liquid chromatography and capillary column gas liquid chromatography (CCGLC).

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Optimization of analytical conditions for the determination of nitrosamines in chlorinated tap water by high performance liquid chromatography (액체크로마토그래피를 이용한 수돗물 중 nitrosamine 화합물 분석의 최적화)

  • Han, Ki-Chan;Kim, He-Kap
    • Analytical Science and Technology
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    • v.23 no.6
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    • pp.551-559
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    • 2010
  • This study was conducted to establish an analytical method for the determination of seven nitrosamines in chlorinated tap water by precolumn derivatization followed by high performance liquid chromatography coupled with fluorescence detection. The derivatization procedure was optimized for denitrosation and dansylation, and then two extraction methods, liquid-liquid extraction (LLE) with dichloromethane and solid phase extraction (SPE), were compared. The SPE method employing the optimized derivation procedure showed higher extraction recovery (54.4-88.7%) and reproducibility (1.9-19.4%) than the LLE method (51.4-87.7% and 4.2-33.3%, respectively). The method detection limits were between 0.5 and 4.4 ng/L. When chlorinated water samples were collected from two treatment plants and ten household taps, and analyzed for nitrosamines, Nnitrosodimethylamine (NDMA) was the major compound found between 26.1 and 112 ng/L.

Determination of Aloesin in Plasma by High-Performance Liquid Chromatography as Fluorescent 9-Anthroyl Derivative

  • Kim, Kyeong-Ho;Lee, Jin-Gyun;Park, Jeong-Hill;Shin, Young-Geun;Lee, Seung-Ki;Cho, Tae-Hyung
    • Archives of Pharmacal Research
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    • v.21 no.6
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    • pp.651-656
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    • 1998
  • A sensitive high-performance liquid chromatographic (HPLC) method for the determination of aloesin in plasma was developed. After solid-phase extraction from plasma and derivatization of aloesin and compound AD-1, which was prepared from aloesin as a internal standard, with 9-anthroylnitrile in the presence of quinuclidine, the derivatives were separated on a Inertsil ODS-3 column using acetonitrile/methanol/water (3:1:6) as a mobile phase, and detected fluorimetrically at 460nm with excitation at 360nm. The detection limit of aloesin was 3.2ng/ml in plasma (S/N=3).

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Synthesis, Characterization and Enhanced Selectivity in RP-HPLC of Polar Carbonyl Group Embedded Poly (Vinyl Octadecanoate) Grafted Stationary Phase by Simple Heterogeneous "Graft from" Technique

  • Rana, Ashequl A.;Karim, Mohammad Mainul;Takafuji, Makoto
    • Bulletin of the Korean Chemical Society
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    • v.32 no.1
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    • pp.77-82
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    • 2011
  • A new high performance liquid chromatograpgy (HPLC) stationary phase that possesses an internal carbonyl functional group is synthesized by heterogeneous "graft from" method. This new stationary phase, poly (vinyl octadecanoate) grafted silica (Sil-2) is then characterized by different physico-chemical methods such as diffuse reflectance infrared fourier transform, suspension state $^1H$ NMR, solid state $^{13}C$ CP/MAS NMR, $^{29}Si$ CP/MAS NMR, elemental analysis and thermogravimetric analysis. Chromatographic properties of Sil-2 were evaluated under reversed phase condition by separating polycyclic aromatic hydrocarbons (PAHs) and comparing the chromatographic results with those on polymeric as well as monomeric octadecylated silica stationary phases.

A Simple and Sensitive Assay for Cefepime in Human Plasma Using High Performance Liquid Chromatography

  • Kim, Young-Sun;Yim, Dong-Seok;Lee, Dong-Gun;Lee, Sang-Bok
    • The Korean Journal of Physiology and Pharmacology
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    • v.7 no.4
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    • pp.247-250
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    • 2003
  • A simple and sensitive assay method was developed for cefepime in human plasma using high performance liquid chromatography (HPLC). Cefepime and cefadroxil (the internal standard) were extracted from heparinized human plasma by simple deproteination with perchloric acid. The extract was injected into an Atlantis dC18 column ($250{\times}4.6$ mm; particle size $5{\mu}m$, Waters) and the column was eluted with methanol and 0.01 M dihydrogen phosphate at pH 3.0 (15:85 v:v) as a mobile phase at a flow rate of 0.7 mL/min. Linearity was confirmed for the range 0.25 to $200{\mu}L/mL$ and the limit of quantitation was $0.25{\mu}L/mL$. The retention times were 10.2 min and 13.4 min for cefepime and cefadroxil, respectively. This method was successfully applied to a pharmacokinetic study of cefepime in plasma from bone marrow transplant patients.

Quantitative Analysis of Microcystins, Cyanobacterial Toxins in Soyang Lake (소양호에서 남조류 독소, 마이크로시틴의 정량 분석)

  • Lee, Jeong Ae;Lee, So Yeong;Pyo, Dong Jin
    • Journal of the Korean Chemical Society
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    • v.46 no.6
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    • pp.535-540
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    • 2002
  • It is very difficult to analyze the microcystins, cyanobacterial toxins quantitatively since it exists in a trace level in lakes. In this paper, two different analytical methods were tried to analyze the microcystins, cyanobacterial toxins quantitatively in water samples collected in Soyang lake. The first method was solid phase extraction method fol-lowed by High Performance Liquid Chromatography(HPLC), and the second method was Enzyme-Linked Immu-nosorbent Assay(ELISA) using the monoclonal antibody of microcystin.

Hydrolysis of Lactose in Whey by the BetavD-Galactosidase (Beta-D-Galactosidase에 의한 유청에 함유된 유당의 가수분해)

  • 최미진;허태련
    • Microbiology and Biotechnology Letters
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    • v.20 no.1
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    • pp.46-52
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    • 1992
  • The optimum condition for the developement of a whey beverage from the concentrated whey was studied. Reverse osmosis system was used to obtain concentrated lactose from cheese whey. The hydrolysis degree of lactose by $\beta$-D-galactosidase was determined using HPLC (high performance liquid chromatography). The order of hydrolysis degree was 1:1, 2:l and 3:l concentrated lactose. It resulted from the concentrated salt which slightly inhibited $\beta$-D-galactosidase with constant enzyme dosage. The optimum condition for enzyme dosage was 2% in non-concentrated lactose, 3% in 2:l and 3% in 3:l concentrated lactose after 4 hours of reaction. When the 3:l concentrated lactose was used, more than 70% was hydrolyzed by 3% enzyme dosage. Furthermore the change of fermented whey by lactic acid bacteria was investigated. Based on the result of sensory test, the most favorable response was obtained at pH 4.2 and titratable acidity of 0.7% about 6 hours of fermentation at $37^{\circ}C$ with 2%: thermophilic starter.

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