Analytical Study of the Antifungal Posaconazole in Raw Material: Quantitative Bioassay, Decomposition Chemical Kinetics, and Degradation Impurities by LC-QTOF-MS.

BACKGROUND: Posaconazole is a triazole antifungal drug that was approved by the U.S. Food and Drug Administration in 2006. No bioassay of it is available in the literature nor official codes for potency determination in bulk. OBJECTIVE: To conduct an analytical study focused on posaconazole in bulk. METHODS: An alternative microbiological assay was validated for drug quantitation, applying agar diffusion technics (3 × 3 design), using Saccharomyces cerevisiae ATCC MYA 1942 as a test microorganism (2% inoculum). An isocratic HPLC-DAD method, with C8 Shim-pack column (250 × 4.6 mm, 5 µm) and methanol-water (75:25 v/v) mobile phase was used for stress stability by photolysis and oxidation, indicating the formation of degradation products, which were investigated by ultra-performance liquid chromatography to quadrupole time-of-flight mass spectrometry. RESULTS: The established conditions for the bioassay were satisfactory. It was linear in the range evaluated (2.5-10.0 µg/mL), as well as precise, accurate, and robust. Stress tests showed drug susceptibility to the factors evaluated (60% of degradation after 120 min). Kinetics curves for photolytic decomposition followed first-order kinetics. From a photolytic and oxidative degraded matrix, three major degradation products were identified as being derivatives with modifications in the piperazine central ring and in the triazole and triazolone side chains, whose mass spectra results were m/z 683 (DP1), m/z 411 (DP2), and m/z 465 (DP3). CONCLUSIONS: The microbiological method was adequately validated and demonstrated to be equivalent to physico-chemical ones. The impurities found are described for the first time in studies with posaconazole raw material. HIGHLIGHTS: A microbiological bioassay was developed for posaconazole, first-order kinetics was determined for photolytic degradation, and structures for new degradation products were suggested.

UV Spectrophotometric method for determination of posaconazole: comparison to HPLC

The aim of this work was to develop a simple, fast and reproducible spectrophotometric method for the analysis of posaconazole in raw material. The established conditions were: methanol as extracting solvent, detection wavelength of 260 nm, Shimadzu double beam spectrophotometer 1800 model with 1 cm quartz cells. Linearity was demonstrated in the concentration range of 5.0 a 25.0 µg/mL (r = 0.9999). Reproducibility and intermediate precision were confirmed by low RSD values (0.49 to 0.82%). Accuracy, evaluated through recovery test, was adequate, with 98.20% of mean recovery. Specificity and robustness were also demonstrated. The mean amount found for samples was 100.82%. The proposed method was considered suitable for the intended purpose, mainly in routine analysis of quality control laboratories. When compared to the previously developed HPLC, no statistical difference was observed, what made the UV spectrophotometric method a reliable alternative.

O objetivo deste trabalho foi desenvolver um método espectrofotométrico simples, rápido e reprodutível para a análise de posaconazol na matéria-prima. As condições estabelecidas foram: metanol como solvente extrator, comprimento de onda de 260 nm e espectrofotômetro de duplo feixe Shimadzu, modelo 1800, com cubetas de quartzo de 1 cm. A linearidade foi demonstrada na faixa de concentração de 5,0 a 25,0 µg/mL (r = 0,9999). A reprodutibilidade e a precisão intermediária foram confirmadas pelos baixos valores de desvio padrão relativo (0,49 a 0,82%). A exatidão, avaliada pelo teste de recuperação, foi adequada, com recuperação média de 98,20%. A especificidade e a robustez também foram demonstradas. O teor médio encontrado nas amostras foi de 100,82%. O método proposto foi considerado adequado, principalmente para a análise de rotina em laboratórios de controle de qualidade. Quando comparado com o método por HPLC, não houve diferença estatística, o que torna o método por espectrofotometria UV uma alternativa segura.