Influence of the peak height distribution on separation performances: Discrimination factor and effective peak capacity

Summary A new index of performance of the chromatographic separation between two adjacent peaks, the discrimination factor, d₀, is defined. It is normalized between 0 and 1 and is directly and easily determined from the chromatogram. It does not depend on any assumption regarding peak shape, except that the peak profiles of individual sample components have a single mode. Its value depends on the relative heights of the two peaks as well as on their separation. The separation power of a chromatographic system is classically measured by its peak capacity, defined on the basis of constant resolution between adjacent peaks. A previously developed statistical theory of the composition of mixtures makes it possible to extend the concept of peak capacity by taking into account the peak height distribution in typical average chromatograms. A new parameter, the effective peak capacity, is defined for this purpose on the basis of a constant discrimination factor between adjacent peaks. It allows to take into account the distribution of peak heights in statistical theories of the evaluation of complex chromatograms and in the measurement of the limit of determination in quantitative analysis. The characteristics of the two new parameters, the discrimination factor and effective peak capacity, are discussed and compared with those of their classical homologs, resolution and peak capacity, in the case of gaussian component peaks of equal widths.     

Implications of the solvent effect in quantitative capillary gas chromatography of minor constituents in mixtures—a study using deuterium labeled and unlabeled benzofluoranthenes

Perdeuterated benzofluoranthenes have slightly shorter retention times than their equivalent unlabeled forms in a DB-5 capillary column. Under column overload conditions, perdeuterated benzofluoranthenes in moderate multifold excesses, which might be encountered in their use as internal standards and carriers for quantitative analysis, are seen to exhibit both normal and reverse solvent effects on their close eluting congeners. In some cases the effects may be used to advantage by knowledgeable analysts, but for the ignorant and unwary the effects can lead to serious errors in identification and quantification.     

An analytical solution of a newly proposed peak broadening equation and extension of polydispersities to higher molecular weight averages in size exclusion chromatography (SEC)

Summary Herein is reported an analytical solution to the peak broadening or peak dispersion/flattening equation based on the recently proposed Instrumental Spreading Shape Function and its application to correction for imperfect resolution (inadequate peak separation and/or excessive peak broadening) for higher molecular weight averages. The relationship of these higher MW averages with the familiar Weight Average and number average molecular weights is also discussed. Criteria for perfect resolution are specified and a true molecular weight calibration curve is accordingly defined.     

Band broadening evaluation by back-and-forth capillary electrophoresis

The purpose of this study is to develop a method for precise evaluation of band broadening in capillary electrophoresis. A capillary electrophoresis system with on-column twin detectors is constructed. Back-and-forth electrophoresis permits zones to make a round trip between the twin detectors placed at distant positions on the capillary. The system is capable of eliminating extra-column effects and discriminating between reversible and irreversible band-broadening mechanisms.     

Unusual peak defocusing in capillary GC on hexakis(2,6-diO-pentyl-3-O-acetyl)-α-cyclodextrin stationary phase

An unusual peak defocusing effect influencing chromatographic performance over a limited range of elution temperatures is described for hexakis(2,6-di-O-pentyl-3-O-acetyl)-α-cyclodextrin stationary phase. Since this phenomenon is likely to be dependent on minor details of the cyclodextrin molecule, full assignment of the ¹H- and ¹³C-NMR-spectra are given.     

Influence of salt and acetonitrile on the capillary zone electrophoresis analysis of imatinib in plasma samples

A simple, sensitive, specific, and cost‐effective analytical methodology was developed for the analysis of human plasma samples spiked with imatinib by CZE with on‐line UV detection in the context of Therapeutic Drug Monitoring. Several analytical conditions such as the ionic strength (I) and the pH of the BGE composed of citric acid and ε‐amino caproic acid were studied in regards of the presence of sodium chloride (NaCl) in plasma samples (1% m/v). Computer simulations (Simul software) were used to confirm the experimental results and to understand imatinib electrophoretic behavior in the presence of NaCl. Furthermore, the advantages of adding ACN to the sample containing NaCl to combine efficient protein precipitation and on‐line CZE stacking of imatinib were demonstrated. LOD and LOQ values of 48 and 191 ng/mL were obtained from plasma sample supernatant after protein precipitation with ACN, which is much lower than mean imatinib plasma level observed for patients treated by imatinib mesylate (about 1000 ng/mL). Good linearity was obtained in the concentration range 191–5000 ng/mL (R² > 0.997). RSD of less than 1.68% and 2.60% (n = 6) for migration times and corrected peak areas, respectively, were observed at the LOQ.     

峰宽法快速测定程序升温脱附活化能

TPD技术^[1]已成为研究固体表面性质的重要手段,本文在总结前人工作的基础上考察了TPD谱峰峰宽和相应参数与脱附活化能(E_d)之间的变化规律,证明用任意峰宽法或半峰宽法测定E_d值有很好的实用性,该法直接运用TPD谱图参数,无需另外作图,故十分快速、简便,与其它方法比较,结果吻合。     

胰岛素反相制备色谱的方法开发

以胰岛素反相制备色谱方法的开发和优化为目标,通过考察色谱保留参数、峰展宽及样品流出曲线的浓度分布等色谱参数,对流动相梯度、色谱填料、载样量等色谱条件进行了优化,并建立了胰岛素制备色谱峰参数的描述方法。结果表明,所建立的方法可快速筛选出最适于胰岛素分离的色谱条件(包括流动相梯度及分离填料),即流动相中的强洗脱溶剂(有机相)需采取缓梯度窄区间的变化条件,筛选出的分离填料需具备峰向两侧展宽且展宽程度较小、样品最高浓度居中分布的特点。将方法用于实际胰岛素粗品的纯化制备,获得了杂质去除效果好、胰岛素纯度高的产品。该法为胰岛素反相色谱纯化制备方法的快速建立提供了指导,具有较强的实用价值,同时为发展大分子化合物的制备色谱方法提供了参考。     

Automatic peak detection in chromatography

Summary A modified derivative of chromatographic signal is proposed for detection of the appearance of a new peak. The method is shown to yield peak areas of good precision even in very noisy systems.This paper is dedicated to Dr. Leslie Ettre on his 60th birthday.     

On the statistical independence of various column contributions to band broadening. Part 1: Second moment contributions from statistically independent,longitudinal diffusion in both phases

The total length-based second moment contribution from longitudinal sample diffusion in both phases on a column, σD , is derived by adding individual partial differential contributions to a partial differential equation accounting for the longitudinal diffusion processes only. Although each diffusion-dispersed sample part is equilibrated between two phases, the resulting σ,D (= 2D _mt _m + 2D _st _s) can be interpreted as the sum of two independent contributions in accordance with the variance addition rule. (D _m and D _s are the mean diffusion coefficients and t _mand t _s the mean residence times of the sample in the mobile and stationary phases, respectively.) The same σD expression is derived from the random walk model of Giddings by treating the diffusional process in each phase as statistically independent of the other processes. Under these conditions the broadening contribution from longitudinal diffusion in the mobile phase is shown to be independent of the velocity profile.