QbD-Driven Development and Validation of Liquid Chromatography Tandem Mass Spectrometric Method for the Quantitation of Sildenafil in Human Plasma.

The present work was employing the Quality by Design approach for the development and validation of a LC-MS-MS method to support the clinical advancement in determination of sildenafil in human plasma using lorazepam as an internal standard. Sample preparation involved solid phase extraction and calibration range observed between 3 and 1,700 ng/mL. The method was systematically optimized by employing Box-Behnken design and used mobile phase flow rate, pH and composition of mobile phase as the critical factors, and assessing the design for retention time and peak area as the responses. A substantial decrease in the variability associated with the method variables was shown in optimization studies and confirmed enhanced method robustness. The present studies revealed that developed method achieves all the regulatory requirements for linearity, accuracy, precision, selectivity, sensitivity and stability for the determination of sildenafil in human plasma. There was not any significant change in the stability of the drug shown by stability studies, performed in human plasma through freeze-thaw cycles, bench-top stability, short-term stability, long-term stability and auto sampler stability. In short, this method shows satisfactory results for the analysis of sildenafil in human plasma and possesses high degree of utility in pharmacokinetic and bioequivalence studies.

Development and validation of LC-MS/MS method for the quantitation of lenalidomide in human plasma using Box-Behnken experimental design.

For the determination of lenalidomide using carbamazepine as an internal standard, an ultra-fast stability indicating LC-MS/MS method was developed, validated and optimized to support clinical advancement. The samples were prepared by solid-phase extraction. The calibration range was 2-1000 ng mL(-1), for which a quadratic regression (1/x(2)) was best fitted. The method was validated and a 3(2) factorial was employed using Box-Behnken experimental design for the validation of robustness. These designs have three factors such as mobile phase composition (X(1)), flow rate (X(2)) and pH (X(3)) while peak area (Y(1)) and retention time (Y(2)) were taken as response. This showed that little changes in mobile phase and flow rate affect the response while pH has no affect. Lenalidomide and carbamazepine were stable in human plasma after five freeze thaw cycles, at room temperature for 23.7 h, bench top stability for 6.4 h. This method competes with all the regulatory requirements for selectivity, sensitivity, precision, accuracy, and stability for the determination of lenalidomide in human plasma, as well as being highly sensitive and effective for the pharmacokinetic and bioequivalence study of lenalidomide.