Method for Gradient Elution in Micro-Flow Liquid Chromatography

This paper describes a new method for gradient elution at low flow rate. By adjusting the solvent composition flowing into a dynamic mixer, we can program the concentration of solvent, B, inside a dynamic mixer to form a time-resolved solvent gradient delivered to the column. The input gradient is related to the output gradient by the equation, B_in = B_out + τ × dB_out/dt, in which τ is the gradient response time constant at specified flow rate, and B_in and B_out are input and output concentration of solvent B, respectively.     

A Simple Device for the Solute Elution from Thin-Layer Chromatographic Plates to High-Performance Liquid Chromatographic or Mass Spectrometric Instrument

JPC – Journal of Planar Chromatography – Modern TLC -     

计算机辅助高效液相色谱梯度洗脱的优化

本文介绍了计算机辅助反相高效液相色谱梯度洗脱优化分离的方法。首先采用混合设计法，通过９个预试验建立目标函数，然后运用双因素（初始流动相组成Ｃ和梯度时间Ｔ）扫描技术，由计算机给出优化分离条件，实验结果和预示完全一致。     

中药指纹图谱的线性梯度分离条件的快速优化方法

提出了一种复杂样品线性梯度分离条件的快速优化方法.先通过4次线性梯度的初始实验,然后利用Excel规划求解工具获得准确的溶质保留方程系数.在此基础上利用网格搜索完成线性梯度的参数优化.整个优化策略通过对中药金银花提取液中各组分的分离得以验证.同时比较了网格搜索、遗传算法、遗传算法与规划求解联用的3种寻优算法.结果表明:3种方法优化结果接近,但运算时间有所差异.     

A Computer Program for the Selection of Gradient Elution in High Performance Liquid Chromatography

Abstract

A computer program is presented for the selection of a gradient mobile phase that will give the same resolution for all the component pairs in the mixture. Each pair is treated as a separate experiment. The computer is used to compile these experiments and to recommend an optimum gradient.     

Quantitative Determination of Polypeptides by Gradient Elution High Pressure Liquid Chromatography

Abstract

High pressure liquid chromatographic (HPLC) methods are described for the quantitative determination of polypeptides in pharmaceutical dosage forms or in synthetic and natural products.

Owing to the addition of an electrolyte for the inactivation of the residual silanol groups present on the modified silica surface, hydrophobic stationary phases were found which proved to be very suitable for the analyses of polypeptides.

Excellent results were obtained with tetramethyl-ammonium containing buffer systems at pH 3 in a gradient of water-methanol. Thus the polypeptides arginine-, lysine vasopressin, oxytocin, ACTH (1–24), ACTH, β-endorphin, glucagon and porcine- and bovine Insulin could be determined with coefficients of variation below 2%.     

A Simple High-performance Liquid Chromatographic Assay for Loracarbef in Human Plasma

Abstract

A simple, rapid and reproducible high-performance liquid chromatographic (HPLC) method for the determination of loracarbef in human plasma has been developed and evaluated. Plasma protein was precipitated with ammonium sulfate. The drug and the internal standard (Cefetamet) were eluted from a μ-bondapak C-18 column with a mobile phase consisting of acetonitrile:methanol:water:glacial acetic acid (2.5:17.5:79.2:0.8%, v/v). The column eluent was monitored at 265 nm. Quantification was achieved by the measurement of the peak-height ratio of the analyte to the internal standard and the limit of quantification for loracarbef in plasma is 0.5 ug/ml. The within-day coefficient of variation (CV) ranged from 2.28% to 3.67%, and between-day CV from 2.38% to 5.59% at three different concentrations. The absolute recoveries ranged from 91.1% to 93.88%, and the relative recoveries from 93.4% to 108% at three different concentrations. Preliminary stability tests showed that loracarbef is stable for at least 5-weeks in human plasma after freezing. The method is applied for the determination of the pharmacokinetic parameters of loracarbef after oral administration to 2 beagle dogs.     

Choice of Model for Estimation of Adsorption Isotherm Parameters in Gradient Elution Preparative Liquid Chromatography

The inverse method is a numerical method for fast estimation of adsorption isotherm parameters directly from a few overloaded elution profiles and it was recently extended to adsorption isotherm acquisition in gradient elution conditions. However, the inverse method in gradient elution is cumbersome due to the complex adsorption isotherm models found in gradient elution. In this case, physicochemically correct adsorption models have very long calculation times. The aim of this study is to investigate the possibility of using a less complex adsorption isotherm model, with fewer adjustable parameters, but with preserved/acceptable predictive abilities. We found that equal or better agreement between experimental and predicted elution profiles could be achieved with less complex models. By being able to select a model with fewer adjustable parameters, the calculation times can be reduced by at least a factor of 10.

     

Simple Liquid Chromatographic Determination of Minocycline in Brain and Plasma

A new high-performance liquid chromatography assay was developed for the determination of minocycline in plasma and brain. A solid–liquid extraction procedure was coupled with a reversed-phase HPLC system. The system requires a mobile phase consisting of acetonitrile:water:perchloric acid (26:74:0.25, v/v/v) adjusted to pH 2.5 with 5 M sodium hydroxide for elution through a RP8 column (250 × 3.0 mm, i.d.) with UV detection set at 350 nm. The method proved to be accurate, precise (RSD < 20%) and linear between 0.15–20 μg mL⁻¹ in plasma and 1–20 μg mg⁻¹ in brain. The method was successfully applied to a blood-brain barrier minocycline transport study.     

High Performance Liquid Chromatographic Procedure for Retinoic Acid in Ophthalmic Solution

Abstract

A fast and simple procedure has been developed for the quantitation of retinoic acid in ophthalmic solutions. the procedure involves the use of reverse phase high performance liquid chromatography with a mobile phase containing an amine modifier. This procedure is specific for retinoic acid in the presence of its degradation products and formulation excipients. the method is linear over a wide range of concentrations.     

Gradient Elution Analysis and Prediction in HPLC

Abstract

Gradient Elution (programming of the carrier solvent composition with time) is increasingly used in HPLC to improve resolution. Mixing effects in the chromatographic system however, cause the actual gradient to be different at the columns than that programmed.

This paper shows how the concepts of residence time distribution and superposition were employed to characterize and predict actual gradients in a Spectra Physics SP8000 HPLC.     

高效液相色谱柱清洗与再生方法

分别对反相、正相、离子交换、体积排阻及亲和等色谱柱的清洗与再生方法加以系统综述.使用适当的方法对受污染的色谱柱进行清洗与再生,可恢复其部分分离能力,延长使用寿命.对硅胶基质反相色谱柱,可选择一系列溶剂,按洗脱能力逐渐增强的顺序依次冲洗,或者使用二甲基甲酰胺、乙酰丙酮、SDS及盐酸胍等强洗脱试剂清洗;硅胶基质正相色谱柱用极性逐渐增强的溶剂系统冲洗;有机聚合物、石墨碳及氧化锆固定相色谱柱,用稀酸、稀碱和有机溶剂结合清洗,可将受污染的色谱柱柱效提高50%以上.     

Determination of Pyridinium Ionic Liquid Cations by Reversed Phase Ion-Pair Chromatography Using Gradient Elution

Four pyridinium ionic liquid cations (N-ethyl-pyridinium, N-butyl-pyridinium, N-butyl-4-methyl-pyridinium, N-hexyl-pyridinium) were separated and determined by reversed phase ion-pair chromatography with ultraviolet-visible detection. The effects of ion-pair reagent, acetonitrile concentration and column temperature on the retention and separation of the cations were evaluated. Then the four pyridinium cations could be separated at baseline within 13 min. The detection limits (S/N = 3) were 0.30–0.70 mg L^?1, and relative standard deviations (n = 5) for peak areas were 0.18–0.58%. The method was applied to analyze surface water with recoveries of 99.5–104.0%, which is accurate, reliable and practical.     

A Simple Liquid Chromatographic Method for the Simultaneous Determination of Antidepressants in Pharmaceutical Preparations

Fluoxetine (FT), fluvoxamine (FX), sertraline (ST) and trazodone (TD) are new type of antidepressants acting as selective serotonin reuptake inhibitors (SSRIs). In structures, they all have chromophore and can be easily monitored by UV absorption spectrophotometry. A simple isocratic high‐performance liquid chromatographic method with ultraviolet detection (215 nm) was developed for the simultaneous quantification of FT, FX, ST and TD. The determination range of the method is over 10.0–400.0 μM for each drug. The detection limits (S/N = 3, injection 20 μL) are about 0.1 μM for TD, 0.2 μM for FT, FX and ST. The relative standard deviation and relative error of the method for intra‐ and inter‐day analyses of FT, FX, ST and TD were all below 3.7%. Application of the method to the analysis of FT, FX or ST in pharmaceutical product proved feasible. The method could be used for the quality control assay of the analytes in bulk and in formulations.     

Comparison of Reverse-Phase High-Performance Liquid Chromatographic Methods for Precolumn-Derivatized Amino Acids

Abstract

A comparison was made among five precolumn derivatization techniques for amino acid analysis using reverse-phase high-performance liquid chromatography (HPLC). All chromatographic analyses were conducted using the same instrumentation and a C₁₈ Ultrasphere ODS column (5 μm, 250 × 4.6 mm). The precolumn derivatization methodologies studied included the formation of OPA (o-phthaldialdehyde), DANSYL (dimethylaminonaphthalenesulphonyl), DABSYL (dimethylaminoazobenzenesulphonyl), PTH (phenylthiohydantoin), and PTC (phenylthiocarbamyl) derivatives. The derivatization procedures were evaluated for simplicity, time required, and derivative stability. HPLC analyses of the amino acid derivatives were compared in terms of resolution, sensitivity, reproducibility, and time of analysis.     

Theory of Gradient Elution Liquid Chromatography with Linear Solvent Strength: Part 1. Migration and Elution Parameters of a Solute Band

Based on the previous theoretical developments most notably by Snyder, this report offers the most complete theoretical framework of gradient elution LC with linear solvent strength (LSS). All statements of the theory are formulated as explicit mathematical expressions. The physics of chromatography in general and of the LSS model in particular were used only to justify the most basic mathematical expressions of the framework. Everything else was obtained by means of verifiable mathematical transformations. The framework was used for derivation of the largest systematic collection of mathematical expressions describing migration and elution parameters of a solute band. Majority of these expressions are new. They include not only the elution parameters of a band, but also previously unknown migration parameters as functions of distance and time traveled by the band. The set of the band parameters in this report was chosen on the basis of the needs for the study of the peak width formation (part 2 of this series) and for detailed study of performance of gradient LC similar to that recently published for temperature-programed GC. As an illustration of the utility of several parameters considered here, a simple way of prediction of a possibility of the reversal of a solute elution order due to the change in the gradient steepness has been found.     

Principle and Potential of Gradient Elution in Planar Chromatography

It was the intention of this paper to show some recent developments in thin layer chroma-togrphy aiming at increasing the separation efficiency by instrumental means and techniquesusing existing separation layers. Gradient elution in the normal phase is a most efficient wayto achieve this goal. Multi-dimensional separations by coupling gradient elution column liquid chromatography inthe reversed phase with AMD gradient elution in normal phase is suitable to achieve newdimensions of separation numbers. Assuming that the N numbers reported for HPLC translateinto separation numbers near 100, by coupling the two kinds of chromatography, separationnumbers around 500 become practically usable.