This study reports the use of retention modeling software for the successful method development of 24 injectable antineoplastic agents. Firstly, a generic screening of several stationary and mobile phases (using various organic modifiers and pH) was achieved. Then, an optimization procedure of mobile phase temperature, gradient profile and mobile phase binary composition was conducted through only 28 real experiments using retention modeling software for data treatment. Finally, the optimized separation was achieved with a mobile phase consisting in 10 mM acetic acid at pH 5.1 (A) and acetonitrile (B). A Waters CORTECS^® T3 column (100 × 2.1 mm, 1.6 μm) operated at 25 °C with a gradient time of 17.5 min (0-51%B) at a flow rate of 0.4 mL/min was used. The prediction offered by the retention model was found to be highly reliable, with an average error lower than 1%. A robustness testing step was also assessed from a virtual experimental design. Success rate and regression coefficient were evaluated without the need to perform any real experiment. The developed LC-MS method was successfully applied to the analysis of pharmaceutical formulations and wiping samples from working environment.

Drug impurity and degradation profiling mean the detection, structure elucidation and quantitative determination of impurities and degradation products in bulk drug materials and pharmaceutical formulations. This is today one of the most important fields of activities in pharmaceutical analysis. The reason for this is that unidentified, potentially toxicimpurities are health hazards, and in order to increase the safety of drug therapy, impurities should be identified and determined by selective methods.

The aim of this review is to characterise the state-of-art in the field of impurity and degradation profiling of drugs based on papers published in the last decade. The separation and determination of impurities and degradants with a known structure are discussed, but emphasis is placed on the structure elucidation and determination of new (unknown) impurities and degradation products by off-line and on-line chromatographic-spectroscopic methods. The analytical aspects of enantiomeric purity of chiral drugs are also discussed.

A generic liquid chromatographic method development workflow was developed and successfully applied to the analysis of phytocannabinoids and Cannabis sativaextracts. Our method development procedure consists in four steps:

i)The screening of primary parameters (i.e. stationary phase nature, organic modifier nature and approximate mobile phase pH) was carried out with a generic gradient on a short narrow bore column, using a system able to accommodate numerous solvents/buffers and columns. Instead of complete peak tracking, the number of peaks which can be separated was considered as a response at this level, to save time.

ii)The optimization of secondary parameters (i.e. gradient conditions, mobile phase temperature and pH within a narrow range) requires only 12 initial experiments and the use of HPLC modeling software for data treatment. It allows to find out the best retention and selectivity for the selected compounds. Peak tracking was performed with a single quadruple mass detector in single ion recording mode, and UV detection (in a broad wavelength range).

iii)The refinement step allows to further adjust column efficiency, by tuning column length and mobile phase flow rate. This can also be done virtually using HPLC modeling software.

iv)The robustness testing step was also evaluated from a virtual experimental design. Success rate and regression coefficients were estimated in about 1 min, without the need to perform any real experiment.

At the end, this method development workflow was performed in less than 4 days and minimizes the costs of the method development in liquid chromatography.

Chromatographic methods are progressing continuously. Increasing sample complexity and safety expectations lead to higher regulatory demands, hence challenges in liquid chromatography analysis are rising, even today, when faster and faster chromatographic systems are extensively employed and become widely accessible for successful method development.

The goal of this study was to investigate the impact of mobile phase influences as important factors of selectivity tuning in method development. This would mitigate mobile phase-related robustness issues throughout the method's lifecycle.

To discover and understand these effects, a new module of chromatographic modeling software DryLab (ver. 4.3.4. beta) was introduced and a special experimental design (DoE) was tested, allowing the simultaneous optimization of solvent-dependent parameters, such as gradient time (t_G), ternary eluent composition (t_C) and pH, requiring 18 input experiments (2 × 3 × 3 = 18).

Additionally, the model creation, using a UPLC system and a narrow bore column (50 × 2.1 mm), the entire experimental work could be finished in 2-3 hours. To demonstrate the applicability of this new design, amlodipine and its related pharmacopoeia impurities (A-H) were subjected to be used in a case study. Predicted vs. Experimental (or Verification) runs showed excellent agreement, average retention time deviations were typically less than 1 s. Modelled robustness testing was also performed, elucidating all important mobile phase and instrument parameters that could influence a method's lifetime performance. Furthermore, as the in silico robustness testing is the least time consuming part of the method development process, it can be used extensively to evaluate robustness even at the very early part in stage 1 of the Method Life Cycle (MLC).

Highlights:

• Proper correlation between logD and relative retention in LC.
• logD calculation is a good basis for starting method screening.
• Drylab software modeling tool to comply analytical QbD requirements in regards to lifecycle principles.
• In silico generation of extended knowledge space possible.
• Extreme high robustness of resulting HPLC method with improved reliability and understanding.