• FOCUS: LIFE SCIENCE
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• 제1호
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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.

• Bulletin of the Korean Chemical Society
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• 제13권1호
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• pp.75-79
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• 1992

Differences in chromatographic properties in RPLC of four brands of cyano bonded silica stationary phases are rationalized in terms of the type and relative strength of the solute-stationary phase interactions, which can be readily inferred from multiple linear regression analyses of retention data for a set of standard compounds on the stationary phases under study based on the linear solvation energy relationships (LSERs). Although four brands of cyano bonded columns studied (CPS-Hypersil, Ultrasphere cyano, Spherisorb-CN and ${\mu}$-Bondapak-CN) have similar bonding density and have been prepared from monofunctional cyanopropylsilane reagents, they possess quite different, relative hydrogen bonding (HB) donor and acceptor strengths. Comparison of the retention behavior on a cyano-bonded silica column with that on an ODS column shows that there are significant differences in the strength of HB interactions between the solute and the stationary phase on the two columns with different functionalities. Information on the differences in the interaction characteristics among brands of the cyano-bonded silica columns and between the ODS and cyano-bonded columns can be utilized to optimize the selectivity for a given separation on these columns.

• FOCUS: LIFE SCIENCE
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• 제1호
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• pp.3.1-3.7
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• 2024

The article discusses the critical role of chromatography in the analysis and purification of proteins in biopharmaceuticals, emphasizing the importance of comprehensive characterization for ensuring their safety and efficacy. It highlights the use of Avantor® ACE® HPLC columns for the separation and purification of proteins, focusing on the analysis of intact proteins using reversed-phase liquid chromatography (RPLC) with fully porous particles. This article also details the application of different mobile phase additives, such as TFA and formic acid, and emphasizes the advantages of using type B ultra-pure silica-based columns for efficiency and peak shape in biomolecule analysis. Additionally, it addresses the challenges of analyzing intact proteins due to slow molecular diffusion and introduces the concept of solid-core (or superficially porous) particles, emphasizing their benefits over traditional porous particles for the analysis of therapeutic proteins. Furthermore, it discusses the development of Avantor® ACE® UltraCore BIO columns, specifically designed for the high-efficiency separation of large biomolecules, such as proteins, and demonstrates their effectiveness in achieving high-resolution separations, even for higher molecular weight proteins like monoclonal antibodies (mAbs). In addition, it underscores the complexity of analyzing and characterizing intact protein biopharmaceuticals, requiring a range of analytical techniques and the use of wide-pore stationary phases, operated at elevated temperatures and with relatively shallow gradients. It highlights the comprehensive range of options offered by Avantor® ACE® wide pore columns, including both fully porous and solid-core particles, bonded with a variety of complementary stationary phase chemistries to optimize selectivity during method development. The use of ultrapure and highly inert base silica is emphasized for enabling the use of lower concentrations of mobile phase modifiers without compromising analyte peak shape, particularly beneficial for LC-MS applications. Then the article concludes by emphasizing the significance of reversed-phase liquid chromatography and its compatibility with mass spectrometry as a valuable tool for the separation and analysis of intact proteins and their closely related variants in biopharmaceuticals.

• Steel and Composite Structures
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• 제7권2호
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• pp.135-160
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• 2007

Recently, CFT column has been well-studied and reported on, because a CFT column has certain superior structural properties as well as good productivity, execution efficiency, and improved rigidity over existing columns. However, CFT column still has problems clearing the capacity evaluation between its steel tube member and high-strength concrete materials. Also, research on concrete has examined numerical values for high-strength concrete filled steel square tube columns (HCFT) to explain transformation performance (M-${\phi}$) when a short-column receives equal flexure-moment from axial stress. Moment-curvature formulas are proposed for HCFT columns based on analytic assumption described in this paper. This study investigated structural properties (capacity, curvature), through a series of experiments for HCFT with key parameters, such as strength of concrete mixed design (58.8 MPa), width-thickness ratio (D/t), buckling length to sectional width ratio (Lk/D) and concrete types (Zeolite, Fly-ash, Silica-fume) under eccentric loads. A comparative analysis executed for the AISC-LRFD, AIJ and Takanori Sato, etc. Design formulas to estimate the axial load (N)-moment (M)-curvature (${\phi}$) are proposed for HCFT columns based on tests results described in this paper.

• Asian Journal of Atmospheric Environment
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• 제11권4호
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• pp.283-299
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• 2017

An analytical method using high-performance liquid chromatography (HPLC) with fluorescence (FL) detection was developed for simultaneously analyzing 10 polycyclic aromatic hydrocarbons (PAHs) and 18 nitro-derivatives of PAHs (NPAHs). The two-dimensional HPLC system consists of an on-line clean-up and reduction for NPAHs in the 1st dimension, and separation of the PAHs and the reduced NPAHs and their FL detection in the 2nd dimension after column-switching. To identify an ideal clean-up column for removing sample matrix that may interfere with detection of the analytes, the characteristics of 8 reversed-phase columns were evaluated. The nitrophenylethyl (NPE)-bonded silica column was selected because of its shorter elution band and larger retention factors of the analytes due to strong dipole-dipole interactions. The amino-substituted PAHs (reduced NPAHs), PAHs and deuterated internal standards were separated on polymeric octadecyl-bonded silica (ODS) columns and by dual-channel detection within 120 min including clean-up and reduction steps. The limits of detection were 0.1-9.2 pg per injection for PAHs and 0.1-140 pg per injection for NPAHs. For validation, the method was applied to analyze crude extracts of fine particulate matter ($PM_{2.5}$) samples and achieved good analytical precision and accuracy. Moreover, the standard reference material (SRM1649b, urban dust) was analyzed by this method and the observed concentrations of PAHs and NPAHs were similar to those in previous reports. Thus, the method developed here-in has the potential to become a standard HPLC-based method, especially for NPAHs.

• 한국독성학회:학술대회논문집
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• 한국독성학회 2006년도 추계학술대회
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• pp.65-74
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• 2006

Modem drug discovery requires rapid pharmacokinetic evaluation of chemically diverse compounds for early candidate selection. This demands the development of analytical methods that offer high-throughput of samples. Naturally, liquid chromatography / tandem mass spectrometry (LC-MS/MS) is choice of the analytical method because of its superior sensitivity and selectivity. As a result of the short analysis time(typically 3-5min) by LC-MS/MS, sample preparation has become the rate- determining step in the whole analytical cycle. Consequently tremendous efforts are being made to speed up and automate this step. In a typical automated 96-well SPE(solid-phase extraction) procedure, plasma samples are transferred to the 96-well SPE plate, internal standard and aqueous buffer solutions are added and then vacuum is applied using the robotic liquid handling system. It takes only 20-90 min to process 96 samples by automated SPE and the analyst is physically occupied for only approximately 10 min. Recently, the ultra-high flow rate liquid chromatography (turbulent-flow chromatography)has sparked a huge interest for rapid and direct quantitation of drugs in plasma. There is no sample preparation except for sample aliquotting, internal standard addition and centrifugation. This type of analysis is achieved by using a small diameter column with a large particle size(30-5O ${\mu}$m) and a high flow rate, typically between 3-5 ml/min. Silica-based monolithic HPLC columns contain a novel chromatographic support in which the traditional particulate packing has been replaced with a single, continuous network (monolith) of pcrous silica. The main advantage of such a network is decreased backpressure due to macropores (2 ${\mu}$m) throughout the network. This allows high flow rates, and hence fast analyses that are unattainable with traditional particulate columns. The reduction of particle diameter in HPLC results in increased column efficiency. use of small particles (<2 urn), however, requires p.essu.es beyond the traditional 6,000 psi of conventional pumping devices. Instrumental development in recent years has resulted in pumping devices capable of handling the requirements of columns packed with small particles. The staggered parallel HPLC system consists of four fully independent binary HPLC pumps, a modified auto sampler, and a series of switching and selector valves all controlled by a single computer program. The system improves sample throughput without sacrificing chromatographic separation or data quality. Sample throughput can be increased nearly four-fold without requiring significant changes in current analytical procedures. The process of Bioanalytical Method Validation is required by the FDA to assess and verify the performance of a chronlatographic method prior to its application in sample analysis. The validation should address the selectivity, linearity, accuracy, precision and stability of the method. This presentation will provide all overview of the work required to accomplish a full validation and show how a chromatographic method is suitable for toxirokinetic sample analysis. A liquid chromatography/tandem mass spectrometry (LC-MS/MS) method developed to quantitate drug levels in dog plasma will be used as an example of tile process.

• Food Science and Biotechnology
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• 제14권2호
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• pp.268-274
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• 2005

Novel antimicrobial substance was isolated from seed coat of Canavalia gladiata by extraction with 75% methanol. Isolation and purification were conducted with solvent fractionation and chromatography on silica gel and sephadex LH-20 columns. Each fraction of antimicrobial activity was tested by paper disc method. Single compound was obtained from the 4th fraction of sephadex LH-20 column chromatography using chloroform/methanol (1:4, v/v), and identified as 3,4,5-trihydroxybenzoic acid methyl ester (methyl gallate) based on HPLC, GC/MS, FT-IR, $^1H$ NMR, and $^{13}C$ NMR analyses. This is the first report describing the presence of methyl gallate in C. gladiata.

• 분석과학
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• 제35권5호
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• pp.189-196
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• 2022

When cationic basic compounds are chromatographed using hydro-organic mobile phase, the presence of anionic free silanols in the silica-based stationary phases results in broad and asymmetrical peaks. The addition of an ionic reagent to the mobile phase prevents analytes from accessing free silanols, improving peak shape. In this study, the chromatographic behavior of salbutamol sulfate as a basic compound was investigated under various conditions, including the use of different columns, mobile phases, and ion-pair reagents such as triethanolamine (TEA) and sodium heptane sulfonate (SHS). The retention and peak shape of chromatograms were both evaluated. The results show that pre-conditioning the column with TEA and including it in the mobile phase can prevent cationic analytes from accessing anionic silanols, resulting in improved peak shape. Furthermore, buffering the mobile phase is an important factor in keeping the pH constant throughout the process. The chosen method was validated in part. This study could be helpful for researchers and analyst to solve such problems with cationic basic components.

• KSBB Journal
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• 제15권5호
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• pp.438-443
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• 2000

맥주의 숙성기간을 단축시키기 위해 네 종류의 고정화용 담체를 활용한 고정화 효모 반응기를 시험하였다. 전발효가 끝난 Green Beer (GB)를 효모를 제거하고 열처리할 경우 처 리온도가 높을수록 전구체인 $\alpha$-acetolactate의 diacety 1로의 전 환율이 낮고 전환속도도 빨라 $70^{\circ}C$이상의 온도에서는 4분이면 충분하였다 GB 중의 산소농도는 전구체에서 DA로의 변환율에 매우 큰 영향을 끼쳤는데 그 농도가 낮을수록 전환율이 낮았으며 0.1 ppm 이하의 농도에서는 거의 대부분의 전 구체가 DA이외의 물칠로 전환되었다. 열처리한 전발효 맥주 를 HAN, G-2, FLO, GDC 담체 column에 $\alpha$, 체류시간 80-150분으로 통과시킬 경우 diacety I 농도를 상품 맥주의 품질로 적합한 0.1 ppm이하로 떨어뜨릴 수 있어서 시험한 담체 모두 맥주 숙성용 효모고정화용 답체로의 실용화 가능 성이 있었다. 이 때 세라믹 담체 column의 경우 GDC 담체 column에 비해 미발효된 잔당의 발효에 의한 새로운 DA 전 구체의 생성이 많았다. 위와 같은 방법으로 생산한 맥주를 공장에서 기존의 방법으로 생산한 맥주와 맛을 비교한 결과 미세한 차이가 있으나 불쾌한 느낌을 주는 이취등은 발견되지 않아 고정화 효모 반응기에 의한 고속 숙성공정의 현장적 용 가능성이 높음을 확인할 수 있었다.

• Computers and Concrete
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• 제19권3호
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• pp.315-323
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• 2017

Within the context of continuum mechanics, inelastic behaviours of constitutive responses are usually modelled by using phenomenological approaches. Elasto-plastic damage modelling is extensively used for concrete material in the case of progressive strength and stiffness deterioration. In this paper, a review of the main features of elasto-plastic damage modelling is presented for uniaxial stress-strain relationship. It has been reported in literature that the influence of Alkali-Silica Reaction (ASR) can lead to severe degradations in the modulus of elasticity and compression strength of the concrete material. In order to incorporate the effects of ASR related degradation, in this paper the constitutive model of concrete is based on the coupled damage-plasticity approach where degradation in concrete properties can be captured by adjusting the yield and damage criteria as well as the hardening moduli related parameters within the model. These parameters are adjusted according to results of concrete behaviour from the literature. The effect of ASR on the dynamic behaviour of a beam and a column are illustrated under moving load and cyclic load cases.