Du Young Choi

Separation of whey proteins by anion-exchange membranes

Three strong anion-exchange membranes (CIM QA, Q100 and HiTrap Q) were investigated for the separation of the major proteins, which were contained in whey, such as α-Lactalbumin, BSA and β-Lactoglobulin. Experiments were performed to determine the optimum mobile phase composition for separating the whey proteins using the standard chemicals of the proteins. The mobile phase was buffer A (20 mM piperazine-HCl pH 6.4) and buffer B (buffer A+1 M NaCl) and the linear gradient elution changes of salt concentration were applied. The standard chemicals of the proteins were used to investigate the optimal mobile phase compositions with the three anion-exchange membranes. From the experimental results, it was found that HiTrap Q was the most effective in separating whey proteins.

Theoretical analysis of chromatographic peak asymmetry and sharpness by the moment method using two peptides

The analyses of peak shapes in chromatography are useful in operating chromatographic system. The asymmetry and sharpness of a chromatographic peak are estimated by the reversed-phase adsorption of two standard peptides (angiotensin II bradykinin) on C18. In this work, the average particle diameters of C18 were 5 and 15 μm, while the pore sizes were 100 and 300 Å. The composition of the mobile phase was 50/50 vol. % of a binary mixture of acetonitrile and water with 0.1% TFA, and the particles were packed in a stainless column (4.6×150 mm). The third and the fourth central movement were calculated from the chromatographic elution curves by moment analysis. The peak asymmetry was determined by two theoretical calculations: the asymmetry factor by elution peak analysis and skewness with moment analysis. The sharpness was estimated by the fourth central moment. In this work, the most acceptable skewness was calculated when the pore size was 300 Å. The large excess was observed on small pore size.

Mobile Phase Composition for Resolving Whey Proteins in Reversed-Phase High Performance Liquid Chromatography

Reversed-phase high-performance liquid chromatography was successfully developed for the simultaneous and rapid separation for the main whey proteins, α-Lactalbumin and β-Lactoglobulin. This method consisted of a linear gradient of the two mobile phases of 0.1% trifluoroacetic acid in water and 0.1% trifluoroacetic acid in acetonitrile. The total run time for this separation was approximately 30 min, and α-Lactalbumin was eluted followed byβ- Lactoglobulin. The injection volume was fixed at 20 μl and the flow rate was 1 ml 1/min. The optimum mobile phase composition and gradient conditions to separate α-Lactalbumin and β-Lactoglobulin (A+B) were experimentally obtained at the 15 μm particle with a pore size of 300 Å on the linear-gradient mode.

Protein binding study of isoflavone, perillyl alcohol and S-ibuprofen by high-performance frontal analysis

High-performance frontal analysis (HPFA) was used for a protein binding study of isoflavones (daidzein, genistin, and genistein), enantiomers of perillyl alcohol and S-ibuprofen to human serum albumin (HSA). The analyses were performed on a Develosil and Inertsil 100-Diol-5 column (10 cm×4.6mm). Sodium phosphate solution (pH 7.4, ionic strength 0.17) was used as the mobile phase at a flow rate of 1 ml/min. To ensure the drug to be eluted as a trapezoidal peak with a plateau, injection volumes were each fixed up the zonal profile with an evident plateau appears. The unbound drug concentration was determined from a plateau height of the plateau region after that experimental data were fitted by Scatchard equation. The binding constants (K) and total binding affinities (nK) of drugs to HSA were calculated, respectively.