Green RP-HPLC method for the estimation of carfilzomib in bulk, protein nanocarriers and human plasma: Application of chemometrics and Monte-Carlo simulations

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2024-12, Vol.1249, p.124350, Article 124350
• Main Authors: Panjwani, Drishti, Patel, Asha, Mishra, Deepak, Patel, Shruti, Patel, Viral, Ram Yadav, Mange, Singh, Bhupinder
• Format: Article
• Language: English
• Subjects: Bioanalytical method; Central-composite-design; Chromatography, High Pressure Liquid - methods; Chromatography, Reverse-Phase - methods; Green Chemistry Technology - methods; Humans; Limit of Detection; Linear Models; Liquid chromatography; Monte Carlo Method; Monte-Carlo Simulations; Nanoparticles - chemistry; Oligopeptides - analysis; Oligopeptides - blood; Oligopeptides - chemistry; Principal Component Analysis; Protein-peptide analysis; Reproducibility of Results
• ISSN: 1570-0232; 1873-376X; 1873-376X
• DOI: 10.1016/j.jchromb.2024.124350

•Chemometrics assisted PCA resulted in appropriate wavelength of quantification.•Factor screening via Placket-Burman design and optimization using CCD.•The method resulted in more than 95% drug recovery in protein NPs and human plasma.•Monte Carlo simulations showed high degree of robustness and variability.•The Green analysis showed high greenness of proposed method than available ones. Carfilzomib is a tetrapeptide epoxyketone that has shown potential clinical outcomes in the treatment of multiple myeloma. However, inaccuracies in quantifying such peptide drug products have arisen due to poor stability, low solubility, time-consuming techniques, complex physicochemical properties, and use of non-green solvents with less recyclability. This provides a substantial urge to develop an ecological and sensitive analytical method for quantifying peptide drugs from matrix formulation and biological samples in early as well as lateral stages of product development in pharma industries. As a result, the study aimed to develop a robust ecological method for estimation of carfilzomib via Green RP-HPLC using analytical quality by design (AQbD) paradigms with specific application in protein nanoparticles and biological matrix. Initially, an appropriate wavelength for quantification of carfilzomib was chosen using principal component analysis (PCA) as a chemometric tool.Risk assessment followed by factor screening studies using 8-factor Placket-Burman Design aided in earmarking critical method parameters (CMPs) affecting critical analytical attributes (CAAs). Further, Central Composite Design (CCD) was employed for design space optimisation to demarcate optimum chromatographic conditions, which were corroborated for robustness using Monte-Carlo simulations. The method was validated as per ICH Q2 (R2), followed by quantifying the greenness of the method using Green Assessment tools. The method optimisation resulted in the optimal chromatographic conditions using Green RP-HPLC. The chromatographic system was equipped with a Phenomenex Aeris Peptide-XC C18 column (150 × 4.6 mm × 5 µm), and the mobile phase was composed of isopropanol:methanol:0.1 M PBS (pH 5.5 adjusted using 0.1 % formic acid) (35:45:20v/v), with a 1 ml/min flow rate at a 210 nm ʎmax. The optimised chromatographic conditions resulted in a short retention time (RT) of 4.95 mins, 0.87 tailing factor (TF), 4,875,122 peak area (PA), and 8995 theoretical plate count (TPC). The method demonstrated linearity in