Characterization and comparison of HPLC columns for gradient elution

Abstract

The type of the stationary phase for reversed-phase liquid chromatography significantly affects the sample polarity range that can be covered using gradients of acetonitrile or of methanol in water. The polarity range available for gradient separations of samples containing compou ncls differing in the lipophilic parts of the molecules can be characterized by"hpophihc capacity", PI, based on the retention of retention standards with regular differences in hpophihty, such as homologous alkylbenzenes. "Standard" PI is defined as a quantity independent of the column dimensions ancl efficiency. It is controlled mainly by the chemistry of the bonded phase ancl by the properties of the support materials, but depends significantly also on the nature of the organic solvent in the mobile phase. The gradient hpophilic capacity ancl the sample polarity range that can be elutecl during a "standardized" gradient run were usecl to compare the graclient-elution performance of more than 20 columns packed with various C8, C18 ancl other bonded phase materials on polymeric, monolithic, non-porous ancl superficially porous supports with various shielding of the residual silanol groups, including "aqueous phase" materials designed for the applications in highly aqueous mobile phases. The "lipophilic capacity" approach can be extended to characterize the suitability of various columns for gradient separations of ohgomers ancl lower polymers, as demonstrated on the example of lower poly(ethylene glycols).

Systematic Structural Characterization of Metabolites in Arabidopsis via Candidate Substrate-Product Pair Networks

“…Plotting the retention time versus the alkyl chain length of all members within a series revealed a linear relationship except for a few very early eluting compounds ( Figure 4D). This elution behavior corresponds exactly with expectations from gradient reversed phase LC (Jandera et al, 2003). Using the parameter estimates of the obtained linear models, the retention times of other members within each glucosinolate series that might have been missed upon chromatogram integration and alignment were calculated.…”

Systematic Structural Characterization of Metabolites in Arabidopsis via Candidate Substrate-Product Pair Networks

“…Plotting the retention time versus the alkyl chain length of all members within a series revealed a linear relationship except for a few very early eluting compounds ( Figure 4D). This elution behavior corresponds exactly with expectations from gradient reversed phase LC (Jandera et al, 2003). Using the parameter estimates of the obtained linear models, the retention times of other members within each glucosinolate series that might have been missed upon chromatogram integration and alignment were calculated.…”

Retention and selectivity tests of silica‐based and metal‐oxide bonded stationary phases for RP‐HPLC

“…The parameters m 1 and a 1 are similar and the parameter p = m 1 /a 1 , is close to unity for various bonded alkyl silica columns tested earlier [41].…”

Peer Reviewed: Characterizing HPLC Stationary Phases

“…The parameters a 0 , a 1 , m 0 , m 1 , p, and q can be used to characterize reversed-phase retention and selectivity. In the alkylbenzene series, the contribution of a repeat methylene group to the slope m 1 and the intercept a 1 when log k versus is plotted is almost identical for various bonded alkyl silica columns and is slightly higher for organic polymer columns (49). The parameter p, which is the ratio of m 1 to a 1 , is close to unity for most columns.…”

Systematic Structural Characterization of Metabolites in Arabidopsis via Candidate Substrate-Product Pair Networks

“…Plotting the retention time versus the alkyl chain length of all members within a series revealed a linear relationship except for a few very early eluting compounds ( Figure 4D). This elution behavior corresponds exactly with expectations from gradient reversed phase LC (Jandera et al, 2003). Using the parameter estimates of the obtained linear models, the retention times of other members within each glucosinolate series that might have been missed upon chromatogram integration and alignment were calculated.…”

Retention and selectivity tests of silica‐based and metal‐oxide bonded stationary phases for RP‐HPLC

“…The parameters m 1 and a 1 are similar and the parameter p = m 1 /a 1 , is close to unity for various bonded alkyl silica columns tested earlier [41].…”

Peer Reviewed: Characterizing HPLC Stationary Phases

“…The parameters a 0 , a 1 , m 0 , m 1 , p, and q can be used to characterize reversed-phase retention and selectivity. In the alkylbenzene series, the contribution of a repeat methylene group to the slope m 1 and the intercept a 1 when log k versus is plotted is almost identical for various bonded alkyl silica columns and is slightly higher for organic polymer columns (49). The parameter p, which is the ratio of m 1 to a 1 , is close to unity for most columns.…”

Systematic Structural Characterization of Metabolites in Arabidopsis via Candidate Substrate-Product Pair Networks

“…Plotting the retention time versus the alkyl chain length of all members within a series revealed a linear relationship except for a few very early eluting compounds ( Figure 4D). This elution behavior corresponds exactly with expectations from gradient reversed phase LC (Jandera et al, 2003). Using the parameter estimates of the obtained linear models, the retention times of other members within each glucosinolate series that might have been missed upon chromatogram integration and alignment were calculated.…”

Retention and selectivity tests of silica‐based and metal‐oxide bonded stationary phases for RP‐HPLC

“…The parameters m 1 and a 1 are similar and the parameter p = m 1 /a 1 , is close to unity for various bonded alkyl silica columns tested earlier [41].…”

Peer Reviewed: Characterizing HPLC Stationary Phases

“…The parameters a 0 , a 1 , m 0 , m 1 , p, and q can be used to characterize reversed-phase retention and selectivity. In the alkylbenzene series, the contribution of a repeat methylene group to the slope m 1 and the intercept a 1 when log k versus is plotted is almost identical for various bonded alkyl silica columns and is slightly higher for organic polymer columns (49). The parameter p, which is the ratio of m 1 to a 1 , is close to unity for most columns.…”