G. Vivó-Truyols

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 COFs. In order to correlate COFs with the probability of deconvolution error, an artificial peak crossing was used to generate 73 different peak arrangements, which were deconvolved using three different methods. SEL(s) exhibited the best correlation, which allowed predicting properly the risk of obtaining inaccurate deconvolutions. The optimisation of a poorly resolved mixture of 16 aromatic compounds by reversed-phase liquid chromatography with methanol-water and acetonitrile-water mobile phases was examined to investigate the differences in performance among the resolution criteria. In situations like these, SEL(s) tends to consider acceptable mobile phase compositions with partial coelution, which permits however the deconvolution with low errors. In contrast, p(s) selects compositions where the resolution of some compounds is sacrificed to enhance the separation of others. Scalar NAS was not so favourable as expected, since it depends on sampling frequency and peak widening. SEL(s) was not affected by these factors.

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 independently to the peak model (single-batch approach). The improvement was especially significant for those chromatograms where the peaks were severely affected by the tails of the preceding compounds. Peak shifts among chromatograms, which are a usual source of non-bilinearity, were modelled in a continuous domain instead of in a discrete way, which avoided some drawbacks associated with latent variable methods. An experimental design involving simulated chromatograms was applied to check the method performance. Five main factors affecting the deconvolution were examined: concentration pattern, chromatographic resolution, number of batches and replicates, and noise level, which were evaluated using first- and second-order figures of merit. The method was also tested on three real samples containing compounds showing different overlap. Four multi-batch deconvolution methods were considered differing in the nature of the processed information and kind of peak matching among chromatograms. In all cases, the multi-batch deconvolution yielded better performance than the single-batch approach.

Towards the optimization of complementary systems in reversed-phase liquid chromatography

SummaryPreviously 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 resolved by use of conventional methods.

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 allows acceptable resolution and longer column lifetime, and its implementation is simpler.