Synthesis of bioactive evodiamine and rutaecarpine analogues under ball milling conditions.

Mechanochemical reactions achieved by processes such as milling and grinding are promising alternatives to traditional solution-based chemistry. This approach not only eliminates the need for large amounts of solvents, thereby reducing waste generation, but also finds applications in chemical and materials synthesis. The focus of this study is on the synthesis of quinazolinone derivatives by ball milling, in particular evodiamine and rutaecarpine analogues. These compounds are of interest due to their diverse bioactivities, including potential anticancer properties. The study examines the reactions carried out under ball milling conditions, emphasizing their efficiency in terms of shorter reaction times and reduced environmental impact compared to conventional methods. The ball milling reaction of evodiamine and rutaecarpine analogues resulted in yields of 63-78% and 22-61%, respectively. In addition, these compounds were tested for their cytotoxic activity, and evodiamine exhibited an IC50 of 0.75 ± 0.04 µg mL-1 against the Ca9-22 cell line. At its core, this research represents a new means to synthesise these compounds, providing a more environmentally friendly and sustainable alternative to traditional approaches.

Kynurenic Acid and Its Analog SZR104 Exhibit Strong Antiinflammatory Effects and Alter the Intracellular Distribution and Methylation Patterns of H3 Histones in Immunochallenged Microglia-Enriched Cultures of Newborn Rat Brains.

Kynurenic acid (KYNA) is implicated in antiinflammatory processes in the brain through several cellular and molecular targets, among which microglia-related mechanisms are of paramount importance. In this study, we describe the effects of KYNA and one of its analogs, the brain-penetrable SZR104 (N-(2-(dimethylamino)ethyl)-3-(morpholinomethyl)-4-hydroxyquinoline-2-carboxamide), on the intracellular distribution and methylation patterns of histone H3 in immunochallenged microglia cultures. Microglia-enriched secondary cultures made from newborn rat forebrains were immunochallenged with lipopolysaccharide (LPS). The protein levels of selected inflammatory markers C-X-C motif chemokine ligand 10 (CXCL10) and C-C motif chemokine receptor 1 (CCR1), histone H3, and posttranslational modifications of histone H3 lys methylation sites (H3K9me3 and H3K36me2, marks typically associated with opposite effects on gene expression) were analyzed using quantitative fluorescent immunocytochemistry and western blots in control or LPS-treated cultures with or without KYNA or SZR104. KYNA and SZR104 reduced levels of the inflammatory marker proteins CXCL10 and CCR1 after LPS-treatment. Moreover, KYNA and SZR104 favorably affected histone methylation patterns as H3K9me3 and H3K36me2 immunoreactivities, and histone H3 protein levels returned toward control values after LPS treatment. The cytoplasmic translocation of H3K9me3 from the nucleus indicated inflammatory distress, a process that could be inhibited by KYNA and SZR104. Thus, KYNA signaling and metabolism, and especially brain-penetrable KYNA analogs such as SZR104, could be key targets in the pathway that connects chromatin structure and epigenetic mechanisms with functional consequences that affect neuroinflammation and perhaps neurodegeneration.

Cinchona-alkaloid-based zwitterionic chiral stationary phases as potential tools for high-performance liquid chromatographic enantioseparation of cationic compounds of pharmaceutical relevance.

Enantiomers of cationic compounds of pharmaceutical relevance, namely tetrahydro-ß-carboline and 1,2,3,4-tetrahydroisoquinoline analogs, were separated by high-performance liquid chromatography. Separations were performed on Cinchona-alkaloid-based zwitterionic ion exchanger type chiral stationary phases applied as cation exchangers using mixtures of methanol and acetonitrile or tetrahydrofuran as bulk solvent components containing triethylammonium acetate or ammonium acetate as organic salt additives. On the zwitterionic ZWIX(+) and ZWIX(-) columns investigated, retention and enantioseparation of the studied basic analytes were influenced by the nature and concentration of the organic components of the mobile phase. The effect of organic salt additives on the retention behavior of the studied analytes can be described by the stoichiometric displacement model related to the counterion concentration. Investigations on the structure-retention relationships were performed applying different mobile phase systems for the two types of cationic analytes. For the thermodynamic characterization, parameters such as changes in standard enthalpy (Δ(ΔH°)), entropy (Δ(ΔS°)), and free energy (Δ(ΔG°)) were calculated on the basis of van't Hoff plots derived from the ln α versus 1/T curves. In most cases, enthalpy-driven enantioseparations were observed, with a consistent dependence of the calculated thermodynamic parameters on the mobile phase composition. Elution sequences of the studied compounds were determined in all cases.

Novel (+)-Neoisopulegol-Based O-Benzyl Derivatives as Antimicrobial Agents.

Discovery of novel antibacterial agents with new structures, which combat pathogens is an urgent task. In this study, a new library of (+)-neoisopulegol-based O-benzyl derivatives of aminodiols and aminotriols was designed and synthesized, and their antimicrobial activity against different bacterial and fungal strains were evaluated. The results showed that this new series of synthetic O-benzyl compounds exhibit potent antimicrobial activity. Di-O-benzyl derivatives showed high activity against Gram-positive bacteria and fungi, but moderate activity against Gram-negative bacteria. Therefore, these compounds may serve a good basis for antibacterial and antifungal drug discovery. Structure-activity relationships were also studied from the aspects of stereochemistry of the O-benzyl group on cyclohexane ring and the substituent effects on the ring system.

Effective Activation by Kynurenic Acid and Its Aminoalkylated Derivatives on M-Type K+ Current.

Kynurenic acid (KYNA, 4-oxoquinoline-2-carboxylic acid), an intermediate of the tryptophan metabolism, has been recognized to exert different neuroactive actions; however, the need of how it or its aminoalkylated amide derivative N-(2-(dimethylamino)ethyl)-3-(morpholinomethyl)-4-oxo-1,4-dihydroquinoline-2-carboxamide (KYNA-A4) exerts any effects on ion currents in excitable cells remains largely unmet. In this study, the investigations of how KYNA and other structurally similar KYNA derivatives have any adjustments on different ionic currents in pituitary GH3 cells and hippocampal mHippoE-14 neurons were performed by patch-clamp technique. KYNA or KYNA-A4 increased the amplitude of M-type K+ current (IK(M)) and concomitantly enhanced the activation time course of the current. The EC50 value required for KYNA- or KYNA-A4 -stimulated IK(M) was yielded to be 18.1 or 6.4 µM, respectively. The presence of KYNA or KYNA-A4 shifted the relationship of normalized IK(M)-conductance versus membrane potential to more depolarized potential with no change in the gating charge of the current. The voltage-dependent hysteretic area of IK(M) elicited by long-lasting triangular ramp pulse was observed in GH3 cells and that was increased during exposure to KYNA or KYNA-A4. In cell-attached current recordings, addition of KYNA raised the open probability of M-type K+ channels, along with increased mean open time of the channel. Cell exposure to KYNA or KYNA-A4 mildly inhibited delayed-rectifying K+ current; however, neither erg-mediated K+ current, hyperpolarization-activated cation current, nor voltage-gated Na+ current in GH3 cells was changed by KYNA or KYNA-A4. Under whole-cell, current-clamp recordings, exposure to KYNA or KYNA-A4 diminished the frequency of spontaneous action potentials; moreover, their reduction in firing frequency was attenuated by linopirdine, yet not by iberiotoxin or apamin. In hippocampal mHippoE-14 neurons, the addition of KYNA also increased the IK(M) amplitude effectively. Taken together, the actions presented herein would be one of the noticeable mechanisms through which they modulate functional activities of excitable cells occurring in vivo.

Novel High Affinity Sigma-1 Receptor Ligands from Minimal Ensemble Docking-Based Virtual Screening.

Sigma-1 receptor (S1R) is an intracellular, multi-functional, ligand operated protein that also acts as a chaperone. It is considered as a pluripotent drug target in several pathologies. The publication of agonist and antagonist bound receptor structures has paved the way for receptor-based in silico drug design. However, recent studies on this subject payed no attention to the structural differences of agonist and antagonist binding. In this work, we have developed a new ensemble docking-based virtual screening protocol utilizing both agonist and antagonist bound S1R structures. This protocol was used to screen our in-house compound library. The S1R binding affinities of the 40 highest ranked compounds were measured in competitive radioligand binding assays and the sigma-2 receptor (S2R) affinities of the best S1R binders were also determined. This way three novel high affinity S1R ligands were identified and one of them exhibited a notable S1R/S2R selectivity.

Bismuth Subnitrate-Catalyzed Markovnikov-Type Alkyne Hydrations under Batch and Continuous Flow Conditions.

Bismuth subnitrate is reported herein as a simple and efficient catalyst for the atom-economical synthesis of methyl ketones via Markovnikov-type alkyne hydration. Besides an effective batch process under reasonably mild conditions, a chemically intensified continuous flow protocol was also developed in a packed-bed system. The applicability of the methodologies was demonstrated through hydration of a diverse set of terminal acetylenes. By simply switching the reaction medium from methanol to methanol-d4, valuable trideuteromethyl ketones were also prepared. Due to the ready availability and nontoxicity of the heterogeneous catalyst, which eliminated the need for any special additives and/or harmful reagents, the presented processes display significant advances in terms of practicality and sustainability.

Pharmacokinetics-Driven Evaluation of the Antioxidant Activity of Curcuminoids and Their Major Reduced Metabolites-A Medicinal Chemistry Approach.

Curcuminoids are the main bioactive components of the well-known Asian spice and traditional medicine turmeric. Curcuminoids have poor chemical stability and bioavailability; in vivo they are rapidly metabolized to a set of bioreduced derivatives and/or glucuronide and sulfate conjugates. The reduced curcuminoid metabolites were also reported to exert various bioactivities in vitro and in vivo. In this work, we aimed to perform a comparative evaluation of curcuminoids and their hydrogenated metabolites from a medicinal chemistry point of view, by determining a set of key pharmacokinetic parameters and evaluating antioxidant potential in relation to such properties.Reduced metabolites were prepared from curcumin and demethoxycurcumin through continuous-flow hydrogenation. As selected pharmacokinetic parameters, kinetic solubility, chemical stability, metabolic stability in human liver microsomes, and parallel artificial membrane permeability assay (PAMPA)-based gastrointestinal and blood-brain barrier permeability were determined. Experimentally determined logP for hydrocurcumins in octanol-water and toluene-water systems provided valuable data on the tendency for intramolecular hydrogen bonding by these compounds. Drug likeness of the compounds were further evaluated by a in silico calculations. Antioxidant properties in diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and oxygen radical absorbance capacity (ORAC) assays were comparatively evaluated through the determination of ligand lipophilic efficiency (LLE). Our results showed dramatically increased water solubility and chemical stability for the reduced metabolites as compared to their corresponding parent compound. Hexahydrocurcumin was found the best candidate for drug development based on a complex pharmacokinetical comparison and high LLE values for its antioxidant properties. Development of tetrahydrocurcumin and tetrahydro-demethoxycurcumin would be limited by their very poor metabolic stability, therefore such an effort would rely on formulations bypassing first-pass metabolism.

Stereoselective synthesis and transformation of pinane-based 2-amino-1,3-diols.

A library of pinane-based 2-amino-1,3-diols was synthesised in a stereoselective manner. Isopinocarveol prepared from (-)-α-pinene was converted into condensed oxazolidin-2-one in two steps by carbamate formation followed by a stereoselective aminohydroxylation process. The relative stereochemistry of the pinane-fused oxazolidin-2-one was determined by 2D NMR and X-ray spectroscopic techniques. The regioisomeric spiro-oxazolidin-2-one was prepared in a similar way starting from the commercially available (1R)-(-)-myrtenol (10). The reduction or alkaline hydrolysis of the oxazolidines, followed by reductive alkylation resulted in primary and secondary 2-amino-1,3-diols, which underwent a regioselective ring closure with formaldehyde or benzaldehyde delivering pinane-condensed oxazolidines. During the preparation of 2-phenyliminooxazolidine, an interesting ring-ring tautomerism was observed in CDCl3.

Diversity-oriented Functionalization of Cyclodienes Through Selective Cycloaddition/Ring-opening/Cross-metathesis Protocols; Transformation of a "Flatland" into Three-dimensional Scaffolds With Stereo- and Regiocontrol.

This article presents selective transformations of some readily available cyclodienes through simple chemical procedures into novel functionalized small-molecular entities. The syntheses hereby described involved selective cycloadditions, followed by ring-opening metathesis of the resulting ß-lactam or isoxazoline derivatives and selective cross-metathesis by differentiation of the olefin bonds on the alkenylated heterocycles. The cross-metathesis transformations have been detailed, which were performed under various experimental conditions with the aim of exploring chemodiscrimination of the olefin bonds and delivering the corresponding functionalized ß-lactam or isoxazoline derivatives.

Sensitivity of Rodent Microglia to Kynurenines in Models of Epilepsy and Inflammation In Vivo and In Vitro: Microglia Activation is Inhibited by Kynurenic Acid and the Synthetic Analogue SZR104.

Kynurenic acid is an endogenous modulator of ionotropic glutamate receptors and a suppressor of the immune system. Since glutamate and microglia are important in the pathogenesis of epilepsy, we investigated the possible action of the synthetic kynurenic acid analogue, SZR104, in epileptic mice and the action of kynurenic acid and SZR104 on the phagocytotic activity of cultured microglia cells. Pilocarpine epilepsy was used to test the effects of SZR104 on morphological microglia transformation, as evaluated through ionized calcium-binding adaptor molecule 1 (Iba1) immunohistochemistry. Microglia-enriched rat secondary cultures were used to investigate phagocytosis of fluorescent microbeads and Iba1 protein synthesis in control and lipopolysaccharide-challenged cultures. SZR104 inhibited microglia transformation following status epilepticus. Kynurenic acid and SZR104 inhibited lipopolysaccharide-stimulated phagocytotic activity of microglia cells. Although kynurenic acid and its analogues proved to be glutamate receptor antagonists, their immunosuppressive action was dominant in epilepsy. The inhibition of phagocytosis in vitro raised the possibility of the inhibition of genes encoding inflammatory cytokines in microglial cells.

Efficient Synthesis of New Fluorinated ß-Amino Acid Enantiomers through Lipase-Catalyzed Hydrolysis.

An efficient and novel enzymatic method has been developed for the synthesis of ß-fluorophenyl-substituted ß-amino acid enantiomers through lipase PSIM (Burkholderia cepasia) catalyzed hydrolysis of racemic ß-amino carboxylic ester hydrochloride salts 3a-e in iPr2O at 45 °C in the presence of Et3N and H2O. Adequate analytical methods were developed for the enantio-separation of racemic ß-amino carboxylic ester hydrochlorides 3a-e and ß-amino acids 2a-e. Preparative-scale resolutions furnished unreacted amino esters (R)-4a-e and product amino acids (S)-5a-e with excellent ee values (≥99%) and good chemical yields (>48%).

N-Acetylation of Amines in Continuous-Flow with Acetonitrile-No Need for Hazardous and Toxic Carboxylic Acid Derivatives.

A continuous-flow acetylation reaction was developed, applying cheap and safe reagent, acetonitrile as acetylation agent and alumina as catalyst. The method developed utilizes milder reagent than those used conventionally. The reaction was tested on various aromatic and aliphatic amines with good conversion. The catalyst showed excellent reusability and a scale-up was also carried out. Furthermore, a drug substance (paracetamol) was also synthesized with good conversion and yield.

Angular Regioselectivity in the Reactions of 2-Thioxopyrimidin-4-ones and Hydrazonoyl Chlorides: Synthesis of Novel Stereoisomeric Octahydro[1,2,4]triazolo[4,3-a]quinazolin-5-ones.

The regioselective synthesis of cis and trans stereoisomers of variously functionalized octahydro[1,2,4]triazolo[4,3-a]quinazolin-5-ones was performed. The 2-thioxopyrimidin-4-ones used in the synthesis reacted with hydrazonoyl chlorides in a regioselective manner to produce the angular regioisomers [1,2,4]triazolo[4,3-a]quinazolin-5-ones rather than the linear isomers [1,2,4]triazolo[4,3-a]quinazolin-5-ones. The synthesis process took place with electronic control. The angular regiochemistry of the products was confirmed by X-ray experiments and two-dimensional NMR studies.

Synthesis of New C-3 Substituted Kynurenic Acid Derivatives.

The application of kynurenic acid (KYNA) as an electron-rich aromatic system in the modified Mannich reaction has been examined. The extension possibility of the reaction was tested by using amines occurring in a number of bioactive products, such as morpholine, piperidine, or N-methylpiperazine and aldehydes of markedly different reactivities, like formaldehyde and benzaldehyde. The influence of substituents attached to position 3 on the aminoalkylation was also investigated. Thus, reactions of 3-carbamoyl-substituted precursors with tertiary amine containing side-chains were also tested to afford new KYNA derivatives with two potential cationic centers. By means of NMR spectroscopic measurements, supported by DFT calculations, the dominant tautomer form of KYNA derivatives was also determined.

Synthesis and Conformational Analysis of Naphthoxazine-Fused Phenanthrene Derivatives.

The synthesis of new phenanthr[9,10-e][1,3]oxazines was achieved by the direct coupling of 9-phenanthrol with cyclic imines in the modified aza-Friedel-Crafts reaction followed by the ring closure of the resulting bifunctional aminophenanthrols with formaldehyde. Aminophenanthrol-type Mannich bases were synthesised and transformed to phenanthr[9,10-e][1,3]oxazines via [4 + 2] cycloaddition. Detailed NMR structural analyses of the new polyheterocycles as well as conformational studies including Density Functional Theory (DFT) modelling were performed. The relative stability of ortho-quinone methides (o-QMs) was calculated, the geometries obtained were compared with the experimentally determined NMR structures, and thereby, the regioselectivity of the reactions has been assigned.

The Antioxidant, Anti-Inflammatory, and Neuroprotective Properties of the Synthetic Chalcone Derivative AN07.

Chalcones belong to a class of biologically active polyphenolic natural products. As a result of their simple chemical nature, they are easily synthesized and show a variety of promising biological activities. 2-Hydroxy-4'-methoxychalcone (AN07) is a synthetic chalcone derivate with potential anti-atherosclerosis effects. In this study, we demonstrated the novel antioxidant, anti-inflammatory, and neuroprotective effects of AN07. In RAW 264.7 macrophages, AN07 attenuated lipopolysaccharide (LPS)-induced elevations in reactive oxygen species (ROS) level and oxidative stress via down-regulating gp91phox expression and stimulating the antioxidant system of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) pathways, which were accompanied by increased glutathione (GSH) levels. Additionally, AN07 attenuated LPS-induced inflammatory factors, including NO, inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and phosphorylated inhibitor of nuclear factor kappa B-alpha (p-IκBα) in RAW 264.7 macrophages. However, the effects of AN07 on promoting nuclear Nrf2 levels and decreasing COX-2 expressions were significantly abrogated by the peroxisome proliferator-activated receptor-γ (PPARγ) antagonist GW9662. In human dopaminergic SH-SY5Y cells treated with or without methylglyoxal (MG), a toxic endogenous by-product of glycolysis, AN07 up-regulated neurotrophic signals including insulin-like growth factor 1 receptor (IGF-1R), p-Akt, p-GSK3ß, glucagon-like peptide 1 receptor (GLP-1R), and brain-derived neurotrophic factor (BDNF). AN07 attenuated MG-induced apoptosis by up-regulating the B-cell lymphoma 2 (Bcl-2) protein and down-regulating the cytosolic expression of cytochrome c. AN07 also attenuated MG-induced neurite damage via down-regulating the Rho-associated protein kinase 2 (ROCK2)/phosphorylated LIM kinase 1 (p-LIMK1) pathway. Moreover, AN07 ameliorated the MG-induced down-regulation of neuroprotective Parkinsonism-associated proteins parkin, pink1, and DJ-1. These findings suggest that AN07 possesses the potentials to be an anti-inflammatory, antioxidant, and neuroprotective agent.

Synthesis of novel fluorinated building blocks via halofluorination and related reactions.

A study exploring halofluorination and fluoroselenation of some cyclic olefins, such as diesters, imides, and lactams with varied functionalization patterns and different structural architectures is described. The synthetic methodologies were based on electrophilic activation through halonium ions of the ring olefin bonds, followed by nucleophilic fluorination with Deoxo-Fluor®. The fluorine-containing products thus obtained were subjected to elimination reactions, yielding various fluorine-containing small-molecular entities.

Cyclodextrin-mediated capillary electrophoresis enantioseparation of dansylated ß-amino acids with bicyclo[2.2.2]octane, bicyclo[3.1.1]heptane and cyclopenta[d][1,2]oxazole core structures.

The present study investigated the separation of bicyclic ß-amino acids with bicyclo[2.2.2]octane, bicyclo[3.1.1]heptane and cyclopenta[d][1,2]oxazole core structures by capillary electrophoresis using native cyclodextrins as well as neutral and charged derivatives as chiral selectors. The amino acids were derivatized with dansyl chloride to provide a UV chromophore. Separations were carried out at 20°C in a 48.5/40 cm, 50 µm fused-silica capillary at an applied voltage of 20 kV. Fifty millimolar sodium phosphate background electrolytes pH 2.5 and 7.2 containing either 5 or 30 mg/mL of the CDs were used. For the majority of the investigated CDs, enantioseparations could only be achieved at pH 2.5 when the analytes are positively charged. Successful enantioseparations as negatively charged analytes at pH 7.2 were only observed for few compounds. In the case of methyl-γ-cyclodextrin, opposite enantiomer migration order was observed in pH 2.5 or 7.2 background electrolytes. Dependence of the enantiomer migration order on the size of the cavity of the cyclodextrins was also found. Furthermore, the degree of methylation of ß-cyclodextrin derivatives affected the migration order of several analyte enantiomers.

HPLC method for the assessment of tryptophan metabolism utilizing separate internal standard for each detector.

The development of a validated method, applicable for the measurement of tryptophan (TRP) and serotonin (5-HT), and that of the neuroprotective branch of the kynurenine pathway from several different biological matrices, including mouse brain, is described. Following the spectral analysis of the metabolites, they were quantified with reversed-phase high-performance liquid chromatography (HPLC), using separate internal standards (ISs) for UV (3-nitro-L-tyrosine) and fluorescent (the newly utilized 4-hydroxyquinazoline-2-carboxylic acid) detectors. With regard to validation parameters, selectivity, linearity, limit of detection, limit of quantification, precision and recovery were determined. Although the linearity ranges were different for the assessed matrices, the correlation coefficient was >0.999 in each case. Furthermore, good intra- and inter-day precision values were obtained with coefficient of variation <5%, and bias <6.5% (except the 5-HT level in brain samples), respectively. The recoveries varied between 82.5% and 116%. The currently developed methods yield opportunities for the assessment of concentration changes in the TRP metabolism from a wide range of biological matrices, therefore they may well be utilized in future clinical and preclinical studies, especially in view that so many metabolites with the application of ISs have not been detected from mouse brain with such a simple HPLC method before.