Separation of selected peptides by capillary electroendoosmotic chromatography using 3 μm reversed-phase bonded silica and mixed-mode phases

The separation of several insect oostatic peptides (IOPs) was achieved by using CEC with a strong-cation-exchange (SCX) stationary phase in the fused-silica capillary column of 75 mm id. The effect of organic modifier, ionic strength, buffer pH, applied voltage, and temperature on peptides' resolution was evaluated. Baseline separation of the studied IOPs was achieved using a mobile phase containing 100 mM pH 2.3 sodium phosphate buffer/ water/ACN (10:20:70 v/v/v). In order to reduce the analysis time, experiments were performed in the short side mode where the stationary phase was packed for 7 cm only. The selection of the experimental parameters strongly influenced the retention time, resolution, and retention factor. An acidic pH was selected in order to positively charge the analyzed peptides, the pI's of which are about 3 in water buffer solutions. A good selectivity and resolution was achieved at pH ,2.8; at higher pH the three parameters decreased due to reduced or even zero charge of peptides. The increase in the ionic strength of the buffer present in the mobile phase caused a decrease in retention factor for all the studied compounds due to the decreased interaction between analytes and stationary phase. Raising the ACN concentration in the mobile phase in the range 40-80% v/v caused an increase in both retention factor, retention time, and resolution due to the hydrophilic interactions of IOPs with free silanols and sulfonic groups of the stationary phase.

Section: Introductionmentioning

confidence: 99%

CEC separation of insect oostatic peptides using a strong‐cation‐exchange stationary phase

Rocco¹,

Aturki²,

D’Orazio³

et al. 2007

“…Several other approaches were introduced to prepare porous polymer monoliths with grafted chemistries for CEC which greatly shortened the time-consuming process of adjusting the monomer composition and optimizing the chromatographic properties [51,52]. During the synthesis, first the beds with well-controlled pore size are prepared, and then the desired surface chemistry is applied onto the surface of the pores of the generic matrices by photoinitiated grafting of suitable polymer chains, necessary to generate EOF [52].…”

Section: Two-step Preparation Of Charged Monolithsmentioning

confidence: 99%

Theoretical and nomenclatural considerations of capillary electrochromatography with monolithic stationary phases

Végvári

Guttman

2006

During the past decade, CEC has been one of the few novel achievements in the field of separation science attracting a wide interest. The technology progress permitted the realization of the long-sought idea to employ an electroosmotically driven flow through the columns improving the separations in terms of both resolution and efficiency. The early practical obstacles related to the use of conventional bead-packed columns have been solved by the introduction of continuous beds, also known as monoliths. Hitherto, various synthesis approaches have been successfully developed producing monolithic beds in situ in capillary columns, sharing similar physical structure built up of tiny particles (in the sub-microm range) that are covalently linked together and to the capillary wall. Parallel with the practical column technology studies, the theory of electrochromatography has been continuously developed, focusing on such basic issues as EOF characterization, separation efficiency, and peak dispersion effects. This review provides a short introduction to the theory of CEC with special attention to monolithic separation beds. The paper also summarizes the latest achievements in CEC and discusses the nomenclature, EOF characteristics, and some specific advantages of monolithic column technology.

“…The retention factor in HPLC can be easily calculated according to the migration time of an analyte and the void time, which is readily available from a chromatogram. Similarly, this definition of retention factor (k*) was applied to CEC according to some authors [14][15][16]:…”

Section: Basic Concepts Of Cecmentioning

confidence: 99%

Polar stationary phases for capillary electrochromatography

Xie

et al. 2004

This review article summarizes the variety of polar stationary phases that have been employed for capillary electrochromatographic separations. Compared with reversedphase stationary phases, the polar alternatives provide a completely different retention selectivity towards polar and charged analytes. Different types of polar stationary phases are reviewed, including the possible retention mechanisms. Electrochromatographic separations of polar solutes, peptides, and basic pharmaceuticals on polar stationary phases are presented.