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Investigation of the analytical performance of aerosol-based detectors in liquid chromatography
thesisposted on 2023-05-27, 10:11 authored by Khandagale, M
A non-discriminating, robust and economical detection system, which can be easily coupled with different modes of separation, is highly desirable in the field of liquid chromatography. While the evaporative light scattering detector (ELSD) and the corona-charged aerosol detector (C-CAD) hold great promise to meet these requirements, the widespread applicability of these techniques has been hindered due to lower sensitivity and solvent dependency of the detection response. This work presents an investigation into different experimental approaches to overcome these limitations, so as to extend the field of applicability of ELSD and C-CAD. Hyphenation of high temperature liquid chromatography (HTLC) using waterrich mobile phases with ELSD / C-CAD is an attractive solution to the solvent dependency limitations of these detectors and also offers a better detection alternative for HTLC. Therefore, experiments were conducted to investigate the effect of HTLC conditions using water-rich mobile phases on the detection response of ELSD and CCAD. Flow-injection studies showed that eluent temperature marginally influenced the detection response. However, in chromatographic separations the response of the ELSD for the same analyte eluted at different retention times was increased up to 5-fold by increasing the separation temperature from 30¬¨‚àûC to 180¬¨‚àûC. Compared to the ELSD, the response of the C-CAD was found to remain relatively unaltered with variation in retention time. This increase in ELSD response was found not to result from the eluent temperature, but rather from compression of the elution band-width at elevated temperatures and hence shorter retention times. The relationship between elution bandwidth and the response mechanism of the ELSD was then explained using logarithmic response curves obtained by flow-injection experiments. Furthermore, it was demonstrated that a temperature gradient could be used to counteract the effects of varying bandwidths associated with isocratic-isothermal separations. Considering the advantages of temperature gradients in attaining elutropic strength comparable to the solvent gradient, the possibility of employing isocratic separations with a combination of temperature and flow-rate variation to achieve uniform detection response of the C-CAD was investigated. Using a flow-injection study, it was demonstrated that the response of the C-CAD remain relatively unaltered with flow-rate variation when used with water-rich eluents. Based on these findings two separation approaches were developed and their utility for C-CAD response normalisation was demonstrated using a mixture of eight analytes. In the first approach, a temperature gradient was applied under isocratic conditions, followed by response enhancement through the post-column addition of organic solvent. In the second approach, flow-rate programming was used to improve the speed of separations performed using isocratic elution coupled with a temperature gradient. The response homogeneity and applicability of these approaches were compared to the inverse solvent gradient technique for quantitative analysis. Good peak area reproducibility (RSD < 15%) and linearity (R2 > 0.994, on a log-scale) over the sample mass range of 0.1 ‚Äö- 10 ˜í¬¿g were achieved. The response deviation across an equi-mass mixture of eight analytes at seven concentration levels was 6-13% compared to 21-39% when a conventional solvent gradient was applied and this response deviation was comparable to that obtained in the inverse gradient solvent compensation approach. The applicability of these approaches for typical pharmaceutical impurity profiling was demonstrated at a concentration of 5 ˜í¬¿g/mL (0.1% of the principal compound). Following the above studies, the applicability of nebuliser gas flow-rate programming and inverse gradient techniques was investigated for improving the performance of the ELSD. The investigations showed that nebuliser gas flow-rate programming could be used to compensate solvent effects; however it caused significant loss in sensitivity and hence has limited applicability for the water rich eluents. Moreover, in inverse gradient experiments, elution bandwidth variability across the separation was found to contribute to response irregularity. This led to the investigation of two-dimensional liquid chromatographic (2-D LC) peak modulation approaches to improve the performance of ELSD. Experiments were conducted to assess the feasibility of elution band-width normalisation by means of post-separation flow-rate modulation using a switching valve. Furthermore, for proof of concept, utility of the switching valve as a peak sampling device to overcome solvent gradient limitations of the ELSD was demonstrated. However, some limitations of this approach were identified, especially in terms of the ability to detect peak segments of low analyte concentration.
Rights statementCopyright 2015 the author Chapter 3 appears to be the equivalent of the post-print version of an article finally published as: M. M. Khandagale, J. P. Hutchinson, G. W. Dicinoski, P. R. Haddad, 2013, Effects of eluent temperature and elution bandwidth on detection response for aerosol-based detectors, J. Chromatogr. A, 1308, 96-103. Chapter 4 appears to be the equivalent of the post-print version of an article finally published as: M. M. Khandagale, E. F. Hilder, R. A. Shellie, P. R. Haddad, 2014, Assessment of the complementarity of temperature and flow-rate for response normalisation of aerosolbased detectors, J. Chromatogr. A, 1356, 180-187.