A novel approach to the qualitative and quantitative assay of HPTLC plates: Second and fourth derivative recording of spectrophotodensitograms

Abstract

Key Words:Thin-layer chromatography, HPTLC Qualitative and quantitative analysis Derivative spectra of analytes SummaryOne of the limitations due to lackof resolution for a given pair of analytes in TLC or HPTLC is the need to optimize the system. In practice this requires time, rerunning of the sample in different developing solvents, and a great deal of expertise on the part of the analyst.In our experience, application of first and second derivative recording techniques toHPTLC facilitates and speeds the whole process, permitting qualitative and quantitative assay of most unresolved spots. Consequently, we have now extended our instrumental capabilities to fourth derivative measurements. For this purpose, we have added a homemade electronic unit in series with the one previously used for first and second order derivatives. Thus, we have been able to evaluate the potential advantages of higher order derivatives for HPTLC analysis of unresolved components in various pharmaceutical products. A comparison of second and fourth order derivative measurements of seriously overlapping HWLC components in a sample of preservatives used in the pharmaceutical industry suggests that the lower order derivatives might be a better choice in view of the higher accuracy and precision of the corresponding data. This is supported by the results of other applications, such as the assay of a commercial colorant, and a syrup formulation. The observed lack of precision of fourth order measurements stems from the fact that although the second and higher order derivatives produce narrower bandwidths, thus contributing to improved resolution, the signal to noise ratio decreases and satellite peak interactions increase, thus rendering correct discrimination of the fine structural detail of overlapping components more difficult.