Rapid and sensitive LC-MS/MS method for the enantioanalysis of verapamil in rat plasma using superficially porous silica isopropyl-cyclofructan 6 chiral stationary phase after SPE: Application to a stereoselective pharmacokinetic study.

A novel liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the rapid and sensitive enantioselective analysis of verapamil (VER) in rat plasma was developed and validated using new superficially porous silica isopropyl-cyclofructan 6 chiral column (LarihcShell-P, LSP). The isocratic mobile phase composed of acetonitrile: trifluoroacetic acid: 10 mM ammonium formate (100 : 0.1 : 0.1, v/v/v) at a flow rate of 0.3 mL/min was applied. Sulpride was utilized as the internal standard (IS). Positive multiple reaction monitoring (MRM) mode was used for mass spectrometry analysis, and the process of analysis was run for 5.2 min. The (S)-(-)- and (R)-(+)-VER enantiomers with the IS were extracted from plasma by using solid-phase extraction (SPE) procedure before the analysis. The C18 cartridge gave good recovery rates for both enantiomers without interference from plasma endogenous. The developed assay was successfully validated following the US-FDA guidelines. The method was linear over concentration ranges of 0.5-500 ng/mL (r2 ≥ 0.997) for each enantiomer (plasma). The lower limits of quantification (LLOQ) for both isomers were 0.5 ng/mL. The intra- and inter-day relative standard deviations (RSD) were less than 8.7 % and the recoveries of (S)-(-)- and (R)-(+)-VER at three spiked levels of 1.5, 250.0 and 450.0 ranged from 92.0%-98.6%. The developed assay was effectively applied in monitoring the stereoselective pharmacokinetic study of VER enantiomers in rat plasma following oral administration of racemic VER. The pharmacokinetic parameters revealed that (S)-(-)-VER demonstrated prominently higher Cmax and AUC values than (R)-(+)-enantiomer. The newly developed approach is the first chiral LC-MS/MS for the quantification of (S)-(-)- and (R)-(+)-VER utilizing superficially porous silica isopropyl-cyclofructan 6 chiral column in rat plasma after SPE.

Enhanced Efficacy of Thiosemicarbazone Derivative-Encapsulated Fibrin Liposomes against Candidiasis in Murine Model.

Candida albicans is the most studied species for the candidiasis infection and is becoming resistant towards existing antifungal drugs. Considering this, in the current study, we developed and characterized a fibrin liposome-based formulation encapsulating a novel thiosemicarbazone derivative, 2C, and evaluated its antifungal efficacy against murine candidiasis. The 2C-containing formulation was prepared by encapsulating 2C within the liposomes (2C-L) that were further encapsulated in the fibrin beads (2C-FL). The in-house synthesized 2C-FLs were spherical with a zeta potential of -34.12 ± 0.3 mV, an entrapment efficiency of 72.6 ± 4.7%, and a loading efficiency of 9.21 ± 2.3%, and they showed a slow and sustained release of 2C. Compared to free 2C, the formulation was non-toxic and exhibited serum stability, increased tissue specificity, and penetration. The 2C-FL formulation had a minimum inhibitory concentration (MIC) value of 4.92 ± 0.76 µg/mL and was able to induce apoptosis and necrosis in C. albicans in vitro. The administration of 2C-FL in C. albicans-infected mice prolonged their survival and antifungal effects when compared with the free 2C. The 2C-FL antifungal therapy significantly reduced the fungal burden and displayed an improved survival rate. In conclusion, the 2C thiosemicarbazone derivative possesses a potent antifungal activity that became more advantageous upon its encapsulation in the fibrin liposome delivery system.

Development and Validation of a Chiral Liquid Chromatographic Assay for Enantiomeric Separation and Quantification of Verapamil in Rat Plasma: Stereoselective Pharmacokinetic Application.

A novel, fast and sensitive enantioselective HPLC assay with a new core-shell isopropyl carbamate cyclofructan 6 (superficially porous particle, SPP) chiral column (LarihcShell-P, LSP) was developed and validated for the enantiomeric separation and quantification of verapamil (VER) in rat plasma. The polar organic mobile phase composed of acetonitrile/methanol/trifluoroacetic acid/triethylamine (98:2:0.05: 0.025, v/v/v/v) and a flow rate of 0.5 mL/min was applied. Fluorescence detection set at excitation/emission wavelengths 280/313 nm was used and the whole analysis process was within 3.5 min, which is 10-fold lower than the previous reported HPLC methods in the literature. Propranolol was selected as the internal standard. The S-(-)- and R-(+)-VER enantiomers with the IS were extracted from rat plasma by utilizing Waters Oasis HLB C18 solid phase extraction cartridges without interference from endogenous compounds. The developed assay was validated following the US-FDA guidelines over the concentration range of 1-450 ng/mL (r2 ≥ 0.997) for each enantiomer (plasma) and the lower limit of quantification was 1 ng/mL for both isomers. The intra- and inter-day precisions were not more than 11.6% and the recoveries of S-(-)- and R-(+)-VER at all quality control levels ranged from 92.3% to 98.2%. The developed approach was successfully applied to the stereoselective pharmacokinetic study of VER enantiomers after oral administration of 10 mg/kg racemic VER to Wistar rats. It was found that S-(-)-VER established higher Cmax and area under the concentration-time curve (AUC) values than the R-(+)-enantiomer. The newly developed approach is the first chiral HPLC for the enantiomeric separation and quantification of verapamil utilizing a core-shell isopropyl carbamate cyclofructan 6 chiral column in rat plasma within 3.5 min after solid phase extraction (SPE).