Interpretive optimisation strategy applied to the isocratic separation of phenols by reversed-phase liquid chromatography with acetonitrile–water and methanol–water mobile phases
An optimisation protocol is presented for the resolution of complex mixtures in isocratic RPLC with binary mobile phases of organic solvent and water, which is based on the prediction of peak position and shape of the individual compounds. A good description of the retention was achieved through the application of statistical weights to the widely used linear or quadratic relationships between the logarithm of the retention factor (log k) and the organic solvent concentration in the mobile phase. The maximisation of the product of peak purities for each compound is shown as a competitive resolution strategy versus the worst value of a selectivity parameter. Peak purities allow one to associ…
Analysis of a solute polarity parameter in reversed-phase liquid chromatography on a linear solvation relationship basis
Abstract A study was made to correlate an overall solute polarity descriptor ( p ) with several molecular parameters: excess molar refraction ( E ), dipolarity/polarizability ( S ), effective hydrogen-bond acidity ( A ) and basicity ( B ), and McGowan volume ( V ), through the linear solvation model ( p = c + eE + sS + aA + bB + vV ). The achieved values of p were introduced in a retention model developed previously for reversed-phase liquid chromatography (RPLC), which describes the retention according to the polarity contributions of solute, mobile phase and stationary phase. The retention behaviour (log k ) of a solute in a given chromatographic system (i.e. column/organic solvent) is a…
Prediction of the retention in reversed-phase liquid chromatography using solute-mobile phase-stationary phase polarity parameters.
Abstract A previously reported algorithm, based on the equation: log k=( log k) 0 +p(P N m −P N s ) , that relates the retention in reversed-phase liquid chromatography with solute (p), mobile phase (PmN) and stationary phase (PsN) relative polarity parameters, is improved. The retention data reported by several authors for different sets of compounds, eluted with acetonitrile–water and methanol–water mixtures, are used to test the algorithm and elaborate a database of p values. The methodology is successfully applied to predict the retention using PmN values calculated as PmN=1.00−(2.13ϕ)/(1+1.42ϕ) for acetonitrile–water and PmN=1.00−(1.33ϕ)/(1+0.47ϕ) for methanol–water, ϕ being the organi…
Considerations on the modelling and optimisation of resolution of ionisable compounds in extended pH-range columns
Abstract The problems associated to the modelling and optimisation of the chromatographic resolution of mixtures involving ionisable solutes at varying pH and acetonitrile content are discussed. Several retention models that separate the contributions of solute, column and stationary phase, were used. The retention was predicted with low errors in large pH domains (2–12), which was an essential requirement to face the optimisation of resolution. The selected mixture was particularly problematic under the viewpoint of resolution, owing to the excessively diverse acid–base behaviour of solutes. This variety led to sudden drops in retention at different pH for each solute, yielding numerous pe…
Determination of the hydrophobicity of organic compounds measured as logPo/w through a new chromatographic method
A new chromatographic method to determine the octanol-water partition coefficient (logP(o/w)) of organic substances is proposed in this paper. This method is based on a previously reported model that relates the retention factor in reversed-phase liquid chromatography with solute (p), mobile phase (P(m)(N)) and stationary phase (P(s)(N)) polarity parameters: logk=(logk)(0)+p(P(m)(N)-P(s)(N)). P(m)(N) values are calculated through expressions that depend only on the organic solvent fraction in the mobile phase. (logk)(0) and P(s)(N) parameters are characteristic of the chromatographic system and are determined from the retention of a selected set of 12 compounds. Then, the p value of a solut…
Hydrophobic and cation exchange mechanisms in the retention of basic compounds in a polymeric column.
A cation exchange retention mechanism concomitant with the well-known hydrophobic partition mechanism in a polymeric column has been observed and investigated. This exchange process is attributed to ionization of some acidic sites present in the polymer column at basic mobile phase pH values. Several drugs of different basicity have been chromatographed on a polymeric PLRP-S column with methanol-water and acetonitrile-water mobile phases. The cation exchange between the protonated basic drug and the buffer cations (Na+, K+ and BuNH4+) is observed at the pH range where the protonated drug and the ionized sites of the column coexist. This process produces a shift of the retention versus pH pl…
Retention-pH profiles of acids and bases in hydrophilic interaction liquid chromatography
Abstract The high proportion of acetonitrile used in many HILIC mobile phases significantly changes the acid-base properties of pH buffers and analytes foreseen from available data in water. In this paper, the recommended stability pH range for chromatographic columns is examined with various acetonitrile/water mixtures, resulting in a significant broadening in the operational pH window with the content of organic solvent. Additionally, the challenge of buffer selection in HILIC is also addressed. Commonly used ammonium acetate shrinks its pH buffering range in acetonitrile-rich mobile phases due to variations in the dissociation constants of the buffer constituents (acetic acid and ammoniu…