HTC-15 - Abstract

Abstract Information :

The analysis of API is required at several stages during the drug development. Monitoring of chemical entities during process development and accelerated stress studies, are just few of the examples where peak identity assignment and quantitation of chemical components is needed in early development stage. However the availability of standards of relevant chemical compounds, such as related impurities, process intermediates etc., at this point of development is limited. The lack of standards prevents quantitation based on calibration curve; additionally peak assignment based on retention time relative to run of the standard is impossible without the respective compounds. Peak assignment could be addressed by monitoring the UV absorption spectra measured with a diode array detector. This approach has severe limitations, for instance the need of a spectra database and most of all it is unsuitable when components co-elute. Therefore it is preferred to hyphenate mass spectrometry to UV detection. In most of cases the detection of the intact mass of the peak is sufficient to confirm an identity, and the availability of the MS spectrum may provide an extra level of confidence in the identity assignment. However this approach does not solve the problem of quantitation. Quantitation without standards is achievable with Charged Aerosol Detector (CAD). With the CAD, the response of non-volatile analytes is highly consistent, provided that the composition of the mobile phase at detector inlet is the same for the all the analytes. This is the case for isocratic elution, or for gradient elution with inverse gradient approach [1]. Thanks to the uniform response, the calibration curve determined with a molecule of choice, for instance the API, will deliver accurate estimation of the amount of the rest of components.

In this work we demonstrate an approach that relies on a multidetector set-up based on LC-UV-CAD-MS. The single quad MS provides m/z values of the peaks to ensure correct peak assignment, and enables detection of co-eluting components. The CAD provides universal quantitation, based on the calibration curve of an available component; in order to ensure that constant mobile phase composition reaches the CAD an inverse gradient will run in parallel and will merge to the analytical flow gradient prior entering the CAD (Figure 1). Moreover the UV detector will complement the data collected by MS and CAD, and acts as valuable tool for method troubleshooting and method transfer.