High Resolution Chromatography and Sensitive Retention: Optimization of the Experimental Conditions for Proteins Extraction by Preparative HPLC

Loai AlJerf, Nuha AlMasri


High performance liquid chromatography (HPLC) is employed with its various elution systems for the fractionation (by isocratic or gradient elution) of peptides and proteins. Lichrosorb Diol (pore size 100 Ǻ) was chosen for normal partition chromatography of proteins. More details about these separations are illustrated in the current research. The chromatographic capacity and its resolution are investigated and the guidelines are widely defined. Reverse-phase (RP) (more suitable with Lichrosorb RP-8) sorbent was partially dissolved during elution with n-propanol (˂ 40 vol. /vol.) and lyophilized during fractionation. An outstanding resolution of these compounds was seen both at pH 4.0 and 7.5 under room temperature and low flow rate at linear gradient of n-propanol. Selective adsorption had been initiated at pH < 4 and peak broadening was observed when salts eliminated from the eluents. It is suggested by the results of this paper, the use of normal phase with Lichrosorb Diol for the isolation of the soluble heterogeneous proteins in extremely elevated concentrations of organic solvents. Consequently, an exceptional resolution, high capacity, and diminutive elution times were verified for peptides and proteins separations.


High performance liquid chromatography; Gradient elution; Proteins; Peak broadening; Adsorption.

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