Analytical validation has fundamental importance in the scope of Good Manufacturing Practice (GMP) for pharmaceutical products since it establishes scientific evidence that an analytical procedure provides reliable results. However, even with validation guidelines available it is very common to observe misunderstandings in the execution of validation and data interpretation. The misguided approaches of validation guidelines, allied with a disregard for the peculiarities of the analytical techniques, the nature of the sample, and the analytical purpose, have significantly contributed to oversights in analytical validation. This work aims to present a critical overview of the validation process in pharmaceutical analysis, addressing relevant aspects of various analytical performance parameters, their different means of accomplishment and limitations in face of the analytical techniques, the nature of the sample, and the analytical purpose. To help in the planning and execution of the validation process, some case studies are discussed, mainly in the area of high-performance liquid chromatography (HPLC).

Perillyl alcohol has been studied in the treatment of cancer disease. However, its high toxicity is a drawback, which can be overcome by its incorporation in nanostructured systems. The aim of this work was to develop and validate a chromatographic method for determination of perillyl alcohol encapsulation efficiency in a polymeric nanoparticles formulation and evaluation of the presence of related degradation products. Perillyl alcohol was subjected to forced conditions of hydrolysis (acidic, alkaline and neutral), oxidation, photolysis and thermal stress, as suggested in the International Conference of Harmonization (ICH) guidelines. The drug showed significant degradation under acidic conditions. The degradation products could be adequately separated on an XBridge C18 column (100 × 2.1 mm, 3.5 µm) using isocratic elution (350 µL min−1) of water/acetonitrile (65 : 35, v/v) at 210 nm. Data from validation studies demonstrated that the method is selective, linear (coefficient of determination (R2) > 0.999) over the range of 20.0-80.0 µg mL−1, precise (relative standard deviation (RSD) < 2.0%), accurate (98.07 to 101.99%) and robust for minor changes. The method was successfully applied to determine the encapsulation efficiency of perillyl alcohol in polymeric nanoparticles, both for product development and for quality control purposes. After nanoparticles production, the presence of degradation products was not observed indicating that the single-emulsion solvent-evaporation technique used does not favor chemical degradation of the drug.

We developed and validated a liquid chromatography tandem mass spectrometry (LC-MS/MS) method to quantify the antimalarials artesunate (ARS) and mefloquine (MFQ) in fixed-dose tablets. The detection was performed by a triple-quadrupole mass spectrometer in multiple reaction monitoring (MRM) in positive ion mode via electrospray ionization. Chromatographic separation was achieved with an XBridge C18 column (50 × 2.1 mm, 5 μm), using isocratic elution (350 μL min-1) of water/acetonitrile/methanol (30:35:35, v/v/v) containing 0.1% formic acid. The method was validated according to the International Conference of Harmonization (ICH) guidelines. The calibration curves obtained for ARS (400 to 600 ng mL-1) and MFQ (800 to 1200 ng mL-1) showed good linearity (r2 > 0.99), precision (relative standard deviation (RSD): ARS < 2.0%; MFQ < 1.9%), and accuracy (recoveries: ARS, 102.4-103.4%; MFQ, 97.4-101.6%), and were stable for 24 h at 8 ºC. The method was successfully applied to commercial tablets, and recoveries of 98.7 ± 4.7% (ARS) and 105.6 ± 3.13% (MFQ). The method developed is a reliable alternative for public quality inspection control with the advantage of tandem mass specificity and speed.