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3-Dimensional Retention Modelling of Gradient Time, Ternary Solvent-Strength and Temperature of the Reversed-phase Gradient Liquid Chromatography of a Complex Mixture of 22 Basic and Neutral Analytes using DryLab®
by Melvin R Euerbya , Gesa Schada , Hans-Jürgen Riegerb , Imre Molnárb ,*
a Hichrom Ltd, 1 The Markham Centre, Station Road, Theale, Reading, Berkshire, RG7 4PE, UK bMolnár-Institute, Schneeglöckchenstr. 47 10407 Berlin, Germany
*Corresponding author - Tel.: +49-30-421-5590, Fax: +49-30-421-55999, Email:
imre.molnar@
molnar-institute.com The present paper describes a multi-factorial optimization of three critical HPLC method parameters, i.e. gradient time (tG –T) plane, which is repeated at three different ternary
2010 ), temperature (T),
and ternary composition (B1:B2) based on twelve experiments for the separation of twenty-two pharmaceutically relevant analytes. Examining the effect of these experimental variables on critical resolution and selectivity was carried out in such a way as to systematically vary all three factors simultaneously. The basic element is a gradient time–temperature (tG
compositions of eluent B between methanol and acetonitrile. The so-defined volume enables the investigation of the critical resolution for a part of the Design Space of a given sample. Multi-dimensional robust regions were successfully defined, graphically depicted and verified. The paper highlights the applicability of this approach for the rapid development of high quality robust LC methodologies.
Keywords: Ternary solvent-strength gradient chromatography; Reversed-phase LC: Computer modelling software; 3-Dimensional model; Robustness of HPLC methods; Method development; Optimization; Quality by Design; Design Space; Validation; QbD; ICH Q8
Introduction
The use of Quality by Design (QbD) and Design Space (DS) principles [1]
is becoming
increasingly popular within the pharmaceutical environment. Regulatory authorities, including the Federal Drug Administration (FDA) and International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH), are actively promoting and demanding the application of these risk-based approaches to drug development in order to ensure a systematic approach in developing analytical methods.
QbD principles, which will now be required for New Drug Applications (NDA), are designed to build in quality from the earliest stage (and
every subsequent stage) of the drug discovery process. The complete information, understanding and transparency of a process relating to risk assessment will now be required for all New Drug Applications – this should speed up the approval processes and, hopefully, eliminate late stage failures.
The move towards QbD in the field of chromatography is a logical consequence of the way in which many HPLC methods have traditionally been developed and validated using a trial and error process. The end result of applying QbD principles to chromatographic method development is an increased understanding of the influence of the chromatographic operating parameters on the analytical measurement (i.e.
chromatographic selectivity, critical resolution, etc), which is achieved through sound science and quality risk management. The final output is the rapid development of chromatographic methods of proven quality and robustness.
L.R.Snyder and his team utilized computer modelling to predict chromatographic retention behaviour and to provide the chromatographer with optimum separations
with a minimum number of input experiments [2]
. In the meantime the new technology has become a commonplace tool for the modern chromatographic method developer [3]
. This
type of HPLC retention prediction and modelling software such as DryLab® (Molnár Institute, Berlin, Germany),
2010
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