N. Chen

The S index in the retention equation in reversed-phase high-performance liquid chromatography

The empirical retention equation log k′ = log k′w − Sϕ in reversed-phase high-performance liquid chromatography (RP-HPLC) was investigated to evaluate the properties of the parameter S. The S index, which is defined as the slope of log k′ versus volume fraction of the organic modifier (ϕ) was systematically examined as a function of bonded phase density, column type and temperature in RP-HPLC. The S index for a particular solute was observed to be nearly constant even when column systems with different C18 packing materials are used. The dependence of log k′ on eluent composition was found to be represented by parallel lines for a given solute for a variety of different stationary phases. The S index remains constant for a given solute despite the prolonged use of C18 column. The results showed that the S index is determined mainly by the interaction between the solute and the mobile phase. It was observed to decrease with the increasing column temperature for non-ionic solutes. Other factors influencing the measurement of the S index are discussed.

Correlation free-solution capillary electrophoresis migration times of small peptides with physicochemical properties

SummaryA series of small peptides containing varying degree of charge and size was used to study the effects of physicochemical properties on migration in free-solution capillary electrophoresis (FSCE). A semiempirical relationship between migration time under acidic conditions and the square root of molecular weight divided by the quantity of the number of the positively ionizable groups has been established. The ionization of the carboxyl terminal group in the polypeptides is negligible under acidic conditions. The relationship developed from this work has been used for the prediction of migration parameters in free solution capillary electrophoresis.

Effects of molecular structure on the S index in the retention equation in reversed-phase high-performance liquid chromatography

Abstract The S index in the retention equation log k′ = log k′w—Sϕ in reversed-phase high-performance liquid chromatography was systematically investigated as the function of molecular structure parameters. The S index, which has been observed to be nearly constant for a specific solute even when column systems with different C18 packing materials are used, was quantitatively correlated with the solvatochromic parameters of the solutes. The coefficients in the correlation of the S index with the solvatochromic parameters of the solutes were investigated and were found to be consistent with the results of using a solvatochromic comparison method. For non-polar compounds, a simplified linear relationship between S and the Van der Waals volume of the solute was observed. For homologues, a linear relationship between S and carbon number was found. Therefore, when other factors remain the same, increasing the size of the solute results in an increase in S whereas increasing the dipolarity or hydrogen bonding ability of the solute will result in a decrease in S.

Influence of column temperature and physico-chemical properties on the electrophoretic behaviour of polyglycine peptides in free-solution capillary electrophoresis

Abstract The changes in electromigration time with column temperature for polyglycine peptides were found to be due to temperature-induced viscosity changes of water. A quantitative linear relationship between the logarithm of migration times (log tm) and the reciprocal of column temperature (1 / T) was derived under a constant electric field strength. The slope of the plot of log tmvs. 1 / T was directly related to the activation energy of diffusion (AED). It was also found that the effect of column temperature on migration times is much more significant under constant-voltage than under constant-current operation. Polyglycine peptides differing only in size were chosen as model molecules for the test of the mobility model. Systematic correlations between tm and the number of glycine amino acids (n) were made. A linear relationship between tm and n0.5 was found for polyglycine peptides under different operating modes. It was observed that the extrapolations of the linear relationships between tm and n0.5 with different column temperatures at constant voltage or with different applied currents at constant temperature cross each other at the same point. The parameters m and m1 in the equation tm = m0 + m1n0.5 were correlated with the column temperature and the operating current. The activation energy of diffusion was also obtained for the plot of log m1 versus 1 / T at constant voltage. Linear relationships between m0 and m1 were found for these peptides which thus result in the intersection point in the plots of tmvs.n0.5.

Electrophoretic Selectivity as a Function of Operating Parameters in Free-Solution Capillary Electrophoretic Separation of Dipeptides

Abstract Free -solution capillary electrophoretic separation of some dipep-tides were carried out under acidic and alkaline conditions. The reproducibility of two migration parameters (migration time and relative migration time) for some dipeptides was examined under various operating conditions. “The electrophoretic selectivity” or relative migration time which is defined as the ratio of the migration time of test component to that of the reference standard shows better reproducibility than migration time. In addition, better reproducibility in migration times was obtained from the capillary previously stored in acidic conditions for a certain period of time. Migration times of a neutral marker (to) and dipeptides (tm) and relative migration times were examined as functions of operating column temperature and applied current (I). A quantitative linear correlation between logarithm of migration time (log to or log tm) and the reciprocal of column temperature (1/T) under constant applied voltage has been d...

The Effect of Column Temperature on the Migration Time of Peptides in Free-Solution Capillary Electrophoresis

Abstract The effect of column temperature on the migration times of peptides in free - solution capillary electrophoresis (FSCE) is discussed. It has been found that there are parallel iinear relationships between the logarithm of the migration time and the reciprocal of column temperature. The changes in migration time with temperature for the peptides are predominantly due to the temperature - induced viscosity changes of water. The elec-trophoretic selectivity remains constant for a specific pair of peptides despite a considerable variation in the migration time for each of the peptides in FSCE.

Peak Identification of the Conjugated Bile Acids in Reversed-Phase Liquid Chromatography by Using the Conjugation Selectivity

Abstract It has been found for the first time that the selectivity between five glycine and taurine conjugated bile acids remains constant despite a considerable variation in retention values for each pair of the compounds in reversed — phase liquid chromatography (RPLC). The dependence of parameter S on log k′w in retention equation log k′ = log k′w—Sϕ results in parallel lines for glycine and taurine conjugates. The selectivity between glycine and taurine conjugates has been utilized for the identification of the conjugated bile acids in RPLC.