Background: Building extensive drug candidate libraries as early in the development pipeline as possible, with high-throughput in vitro absorption, distribution, metabolism, and excretion (ADME) profiling, is crucial for the selection of lead compounds to guide subsequent research and production phases. Traditionally, the analysis of metabolic stability assays heavily relies on high-throughput LC-MS/MS (liquid chromatography tandem mass spectrometry) techniques to meet with the lead profiling demands. Laser-assisted rapid evaporative ionization mass spectrometry (LA-REIMS) is a quick and efficient technique for characterizing complex biological samples without laborious sample preparation. Objective: In this study, using an automated LA-REIMS well plate reader, achieving an 8 s per sample measurement time, the oxidative metabolic stability of active drug agents was assessed using biomimetic metalloporphyrin-based oxidative model reactions. Results: The results obtained using the novel LA-REIMS-based protocol were compared to and corroborated by those obtained using conventional HPLC-UV-MS (high performance liquid chromatography with ultra-violet detection coupled with mass spectrometry) measurements. Conclusions: LA-REIMS emerges as a promising technique, demonstrating potential suitability for semi-quantitative high-throughput metabolic stability in an optimized solvent environment.

Motivated by the in vivo anxiolytic activity of previously described 1,2,3-benzothiadiazine 1,1-dioxides, we aimed at elaborating a synthetic procedure for the preparation of their pyrrole-fused counterparts, 2,9-dihydro[1,2,3]thiadiazino[5,6-g]indole 1,1-dioxide derivatives. The simple and versatile process led, via Fischer indole cyclization of the corresponding hydrazones, to a wide structural variety of new tri-, tetra- and pentacyclic ring systems. The structural characterization of (E)- and (Z)-hydrazones was supported by 2D NMR techniques, while that of the target compounds by single-crystal X-ray measurements. The hydrazone intermediates and the new title compounds were subjected to a physicochemical and early ADME characterization study, in the framework of which logP, pK_a and logk values were calculated. Following that, kinetic solubility and in vitro gastrointestinal membrane-specific permeability measurements were carried out to assess the lead-likeness of the compounds. Subsequently, the metabolic stability of the most promising derivatives was also determined using human liver microsomes.

It is an utmost importance to characterize drug candidates as early in the development pipeline as possible. Appropriate application of the emerging high throughput analytical techniques unlocks the opportunities to obtain experimental data for in vitro absorption, distribution, metabolism and elimination (ADME) profiling of large compound libraries which is crucial for selection of lead compounds. In an usual scenario, only one drug candidate will be selected from thousands of early compounds based on several stability and screening results. A high to medium throughput screening approaches could significantly accelerates the subsequent research and production phases. Traditionally, the analysis of metabolic stability assays heavily relies on high throughput LC-MS/MS techniques to meet with the lead profiling demands. In this study presented an innovative experimental approach that can offer a refreshing alternative. Laser Assisted Rapid Evaporative Ionization Mass Spectrometry (LA-REIMS) is a quick and efficient method for characterizing complex biological samples without laborious sample preparation and chemical separation. Therefore, using an automated LA-REIMS well plate reader as fast as 8 seconds per sample, we assessed the oxidative metabolic stability of active agents using biomimetic metalloporphyrin-based oxidative model reactions at various pH for commercially available drug substances and compared stability measurements with traditional HPLC-DAD-MS results.

Background/Objectives: Stereochemistry can substantially influence the biological activity and pharmaceutical properties of small molecules. This study investigated the stereochemistry-dependent biological activity and chiral recognition of novel naphthylethyl thiourea enantiomers using an integrated biological, chromatographic, and computational approach. Methods: Ten pairs of naphthylethyl thiourea enantiomers were synthesized from enantiomerically pure precursors and characterized. Antiproliferative and antibacterial activity, chiral HPLC behavior on polysaccharide- and protein-based stationary phases, molecular docking, and multivariate relationships were evaluated. Results: Target-dependent enantioselectivity and substituent-dependent activity patterns were observed. All enantiomeric pairs were chromatographically distinguished under at least one condition, and selected compounds showed selector- and mobile-phase-dependent changes in elution order. AGP-based chromatography showed stereoselective recognition for a subset of compounds, while computational analyses provided complementary support for the experimental trends. Conclusions: Stereochemistry and substitution jointly influence biological and chromatographic behavior in this thiourea series. Integrated biological, chromatographic, and computational profiling provides a useful framework for early enantiomer-resolved developability assessment.