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.

1. S002-333 [(2-(4'-methoxy-benzenesulfonyl)-2,3,4,9-tetrahydro-1H-pyrido (3,4-b) indole-3-carboxylic acid amide)] is a novel and potent antithrombotic active agent. The present work investigates the pharmacokinetics, bioavailability, dose proportionality and permeability of the racemate, S002-333 in male New Zealand White (NZW) rabbits. 2. Rabbits were administered single intravenous (i.v.) (2 mg/kg) and three oral doses of 10, 20 and 40 mg/kg of S002-333, respectively, at different occasions to evaluate dose proportionality. Serial blood samples were collected and analyzed by a liquid chromatography tandem mass spectrometry (LC-MS/MS) method. Since S002-333 is a racemate consisting of S004-1032 (R) and S007-1558 (S), same samples were analyzed using a chiralcel column so as to evaluate the respective enantiomers. 3. The peak plasma concentration, after oral administration, occurred at ∼10 h post-dose. The clearance (CL) and volume of distribution (Vd) after i.v. dose were found to be 3.05 ± 0.09 l/h/kg and 6.73 ± 1.16 l/kg, respectively. The absolute oral bioavailability of S002-333 was 16.32%, whereas it was 6.62 and 5.90% for R- and S-enantiomers, respectively. The absolute bioavailability of 10, 20 and 40 mg/kg doses were found to be 27.91, 14.39 and 16.91%, respectively. The PAMPA (parallel artificial membrane permeability assay) assay shows that S002-333 has a low-passive permeability at gastric and intestinal environment. 4. In conclusion, S002-333 has low-passive permeability, low CL and large Vd. The R-enantiomer has a "slightly" greater bioavailability than the S-enantiomer.

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Preclinical Research & Development Withanolide A (WA), a steroidal lactone is a major bioactive constituent of Withania somnifera (L.) with remarkable neuropharmacological activity. In this study, we investigated the permeability, plasma protein binding (PPB), blood partitioning, intravenous (i.v.), and oral pharmacokinetics as well as i.v. tissue distribution (TD) of pure WA in a rat model. The PPB, RBCs partitioning, and permeability of WA were determined by Ultra Performance Liquid Chromatography (UPLC) method. However, the pharmacokinetics and TD of WA were evaluated by validated and sensitive liquid chromatography coupled mass spectrometry (LC‐ESI‐MS/MS) method. The PPB and permeability of WA were determined by equilibrium dialysis and parallel artificial membrane permeability assay method, respectively. The results demonstrated that WA has high PPB and passive permeability. Furthermore, WA was found to have fast equilibration between RBCs and plasma. Following i.v. (2 mg/kg) and per‐oral (25 mg/kg) administration of WA, the max concentration (Cmax) in plasma was found as 85.53 ± 6.54 and 48.04 ±5.78 ng/mL, respectively. The TD study results indicated that WA has a rapid and wide TD. The maximum concentration in various tissues was found in following order: Clung > Cliver > Ckidney ≈ Cspleen > Cheart > Cbrain. The preclinical in vitro, as well as pharmacokinetics and TD results, are anticipated to support the future preclinical and clinical application of WA.

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