0000000000342135
AUTHOR
G. Vivó-truyols
Robust interpretive optimisation in high-performance liquid chromatography considering uncertainties in peak position.
In the context of interpretive chromatographic optimisation, robustness is usually calculated by introducing deliberated shifts in the nominal optimal conditions and evaluating their effects on the monitored objective function, mimicking thus the experimental procedures used in method validation. However, such strategy ignores a major source of error: the uncertainties associated to the modelling step, that may give rise to deceiving results when conditions that were expected to yield baseline separation are reproduced in the chromatograph. Two approaches, based on the peak purity concept, are here proposed to evaluate the robustness of the objective function under the perspective of measur…
Peak deconvolution in one-dimensional chromatography using a two-way data approach.
A deconvolution methodology for overlapped chromatographic signals is proposed. Several single-wavelength chromatograms of binary mixtures, obtained in different runs at diverse concentration ratios of the individual components, were simultaneously processed (multi-batch approach), after being arranged as two-way data. The chromatograms were modelled as linear combinations of forced peak profiles according to a polynomially modified Gaussian equation. The fitting was performed with a previously reported hybrid genetic algorithm with local search, leaving all model parameters free. The approach yielded more accurate solutions than those found when each experimental chromatogram was fitted in…
Error analysis and performance of different retention models in the transference of data from/to isocratic/gradient elution.
The transferability of retention data among isocratic and gradient RPLC elution modes is studied. For this purpose, 16 beta-blockers were chromatographed under both isocratic and gradient elution with acetonitrile-water mobile phases. Taking into account the elution mode where the experimental data come from, and the mode where the retention should be predicted, the following combinations are possible: isocratic predictions from (i) isocratic or (ii) gradient experimental designs; and gradient predictions from (iii) isocratic or (iv) gradient data. Each of these possibilities was checked using three retention models that relate the logarithm of the retention factor: (a) linearly and (b) qua…
Estimation of significant solvent concentration ranges and its application to the enhancement of the accuracy of gradient predictions.
Abstract The solvent concentration range actually useful for gradient predictions is significantly narrower than the total range scanned in a gradient run. This range, called “solvent informative range” (SIR), if known with the highest accuracy, allows to predict gradient retention times ( t g ) with minimal error. The small size of the SIR supports the application of the linear solvent strength theory (LSST). Furthermore, LSST allows a closed-form solution to the integral required to predict gradient retention times, which eliminates numerical integration, needed with other retention models. A methodology that calculates the SIR by applying error analysis, and uses it to improve the accura…
Towards the optimization of complementary systems in reversed-phase liquid chromatography
Previously reported optimization methodology, which seeks complementary mobile phases (CMP) in isocratic chromatography, has been extended to include more than one system simultaneously (i.e. more than one organic solvent and/or column). In the literature the benefits of complementarity are not usually fully exploited—few working conditions giving rise to interactions as different as possible are examined, without developing a fully linked optimization. The proposed approach is compared critically with use of a single mobile phase or CMP which consider one system only. The strategy greatly expands the capability of isocratic chromatography in the analysis of complex samples that cannot be r…
Limits of multi-linear gradient optimisation in reversed-phase liquid chromatography
Abstract The concept of limiting peak purity was applied to quantify the degree of completion of the separation capability of a chromatographic system using multi-linear gradients. The objective was to check whether the complexity of a gradient program deserves be increased to enhance resolution by inserting more linear segments, or on the contrary, no significant improvements can be expected under more complex gradients. A set of 19 isoindole derivatives of primary amino acids was selected to test the performance of isocratic, single linear and multi-linear gradients. Accurate simulated chromatograms were obtained via numerical integration of the general equation of gradient elution, using…
Separation of Proteic Primary Amino Acids under Several Reversed‐Phase Liquid Chromatographic Conditions
Abstract The reversed‐phase liquid chromatographic (RPLC) analysis of proteic primary amino acids with acetonitrile‐water, using pre‐column derivatisation with o‐phthalaldehyde (OPA) and N‐acetylcysteine (NAC), was compared with RPLC modes using trifluoroacetic acid or pentadecafluorooctanoic acid and evaporative light‐scattering detection, or sodium dodecyl sulphate micelles with pre‐ and post‐column derivatisation. The importance of column lifetime, risk of potential damages in the instrumentation, retention and resolution, was considered. Among the assayed approaches, the best is still aqueous‐organic RPLC with pre‐column derivatisation. It not only yields the most reliable results, but …
Net analyte signal as a deconvolution-oriented resolution criterion in the optimisation of chromatographic techniques
The performance of two multivariate calibration measurements, multivariate selectivity (SEL(s)) and scalar net analyte signal (scalar NAS), as chromatographic objective functions (COFs), was investigated. Since both assessments are straightforwardly related to the quantification of analytes in the presence of interferents, they were expected to confer new features in the optimisation of compound resolution, not present in conventional assessments. These capabilities are especially interesting in situations of low resolution, where peak deconvolution becomes an attractive alternative. For comparison purposes, chromatographic resolution (R(s)) and peak purity (p(s)) were used as reference COF…
A comparative study of the performance of acetonitrile and methanol in the multi-linear gradient separation of proteic primary amino acids
The performance of the separation of proteic primary amino acids using multi-linear gradients of acetonitrile and methanol was studied under an experimental-design basis, using an Inertsil ODS-3 column and pre-column derivatization with o-phthaldialdehyde (OPA) and N-acetyl-l-cysteine (NAC). Elution strength, peak properties, resolution, and analysis time, were examined. The optimal separation was established through modeling, using information obtained from isocratic data. By optimizing the separation with gradients of increasing complexity, acceptable resolution was possible, being glycine/threonine the critical pair. Multi-criteria decision-making (Derringer desirabilities) was applied t…
Enhancement of retention predictions in reversed-phase liquid chromatography using reference compounds
Abstract The use of reference compounds to correct errors associated with the preparation of mobile phase is studied, in order to enhance modelling of retention for optimisation purposes in reversed-phase liquid chromatography. The method fits individual retention models for a set of reference compounds, which are present in all injections. In second step, mobile-phase composition is corrected for each chromatogram, according to the fitted models of the reference compounds. The enhanced precision provided by the large number of replicates for the reference compounds is partially propagated to the retention models of the analytes. The approach is assayed in the separation of amino acids, aft…
Enhanced calculation of optimal gradient programs in reversed-phase liquid chromatography
Abstract The resolution of a mixture of 16 β-blockers under gradient elution was optimised using both isocratic and gradient training sets, with a reversed-phase column and acetonitrile–water eluents. Error theory was applied to measure the information extracted from different gradient experimental designs. This allows checking the expected accuracy when gradient predictions exceed the initial solvent concentrations tested in the training set. This work applies the results on modelling found in a previous study [J. Chromatogr. A 1018 (2003) 169] where the performance of several retention models was compared. Enhanced retention predictions were applied to the optimisation of gradient program…
Automatic program for peak detection and deconvolution of multi-overlapped chromatographic signals
Several interlinked algorithms for peak deconvolution by non-linear regression are presented. These procedures, together with the peak detection methods outlined in Part I, have allowed the implementation of an automatic method able to process multi-overlapped signals, requiring little user interaction. A criterion based on the evaluation of the multivariate selectivity of the chromatographic signal is used to auto-select the most efficient deconvolution procedure for each chromatographic situation. In this way, non-optimal local solutions are avoided in cases of high overlap, and short computation times are obtained in situations of high resolution. A new algorithm, fitting both the origin…