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Chromatographic behavior of novel chiral stationary phases with bonded selectors based on Cinchona alkaloids modified with dipeptides was studied using dipeptides as probe molecules. Buffer-free and salt containing hydro-organic solutions were used as the mobile phases. The selectors exhibit pseudoenantiomeric behavior with respect to the L/D or LL/DD enantiomers and do not behave so with respect to the LD/DL enantiomers. The alkaloid part of the selectors is the driver of enantioselectivity, while the dipeptide substituent plays a modulating role. The quinidine-based selectors demonstrate stronger adsorption affinity and higher enantioselectivity as compared to the quinine-based selectors. The dipeptide analytes containing a glycyl fragment are weaker retained and their enantiomers are worse separated comparing to dipeptides with both units being larger amino acids. Moreover, a phenyl group in the structure of a dipeptide analyte facilitates enantioseparation. The effect of the mobile phase composition on retention depends on the hydrophobicity of an analyte. Hydrophobic dipeptides are better eluted by methanol-rich solvents, hydrophilic dipeptides are better eluted with water-rich solvents, and dipeptides with an intermediate hydrophobicity demonstrate a U-shaped or more complicated dependence of the retention factor on the percentage of methanol. Even a small buffer addition to the mobile phase decreases retention, but the ion-exchange mechanism was not confirmed. The effect of an electrolyte is rather due to the shielding of the charged groups of the selector reducing thereby electrostatic interaction between the selector and analyte. Efficiency of the novel columns is comparable to that of other brush-type chiral columns, the highest achieved number of the theoretical plates per 1 m varying between 30,000 and 40,000.

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Chiral resolution of polar organic compounds such as amino acids and peptides represents an important chromatographic task due to increasing significance of natural species, which play important signaling and regulatory roles in the living organisms. Despite the number of available chiral stationary phases, this task remains challenging, since not many of the commercially available systems are capable to resolve non-derivatized zwitterionic species. In this study, we present a target-oriented design of a new class of chiral selectors. Pursuing the goal to separate amino acids, and especially short peptides, we have combined Cinchona alkaloids - quinine and quinidine - with three different biogenic dipeptides. We have synthesized six different chiral stationary phases, with selector loading of ∼200 µmol g-1, and tested their chiral recognition capabilities for acidic, basic and zwitterionic analytes using various mobile phases. We have observed that all chiral stationary phases retain the chiral anion exchange capability known for commercially available Cinchona-based columns leading to baseline or partial resolution of six out of ten analytes. The performance in chiral resolution of basic analytes is not optimum due to the weak cation exchange character of the peptidic residue. However, we report on encouraging results in the chiral resolution of short peptides, for which, depending on their structure, we see the chiral resolution of up to three stereoisomers (from four possible) in a preliminary screening.

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From 2000, our two research groups independently and simultaneously designed and developed a novel family of electrophilic fluorinating reagents based on the use of Cinchona alkaloids. The chiral N-fluoro ammonium salts demonstrated the highest efficiency compared to prior art in enantioselective electrophilic fluorination for a wide range of substrates. In this account, we tell our respective stories, how the same idea germinated in our laboratories, the characterization of the chiral reagents, the use in stoichiometric quantity then the development of a catalytic version, the application to the synthesis of chiral fluorinated molecules of pharmaceutical interest, and finally the exploitation of our reagents by other teams and for other applications.

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BACKGROUND: Glioblastoma multiforme (GBM) is a lethal form of central nervous system cancer with a lack of efficient therapy options. Aggressiveness and invasiveness of the GBM result in poor prognosis and low overall survival. Therefore, the necessity to develop new anti-carcinogenic agents in GBM treatment is still a priority for researchers. Ion channels are one of the primary regulators of physiological homeostasis with additional critical roles in many essential biological processes related to cancer, such as invasion and metastasis. A multi-channel blocker, hydroquinidine (HQ), is currently in use to treat short-QT and Brugada arrhythmia syndromes. OBJECTIVE: The objective of the study was to examine the alterations in survival, clonogenicity, migration, tumorigenicity, proliferation, apoptosis, and gene expression profile of temozolomide (TMZ)-sensitive and TMZ-resistant GBM cells upon HQ treatment. METHODS: The possible anti-neoplastic activity of HQ on GBM cells was investigated by several widely applied cell culture methods. The IC50 values were determined using the MTT assay. Upon HQ treatment, the clonogenicity and migration capacity of cells were evaluated via colony-formation and wound healing assay, respectively. For antiproliferative and apoptotic effects, EdU and CFSE, and Annexin-V labeling were applied. Tumorigenicity level was depicted by employing soft agar assay. The expression level of multiple genes functioning in the cell cycle and apoptosis- related processes was checked utilizing qPCR. RESULTS: A significant anti-carcinogenic effect of HQ on TMZ-sensitive and -resistant GBM cells characterized by the increased apoptosis and decreased proliferation rate was revealed due to the altered gene expression profile related to cell cycle and cell death. CONCLUSION: In this study, the anti-carcinogenic effect of HQ has been demonstrated for the first time. Our data suggest the possible utilization of HQ to suppress the growth of GBM cells. Further studies on GBM-bearing animal models are required to assess its therapeutic potential in GBM treatment.

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Organocatalysis is a very useful tool for the asymmetric synthesis of biologically or pharmacologically active compounds because it avoids the use of noxious metals, which are difficult to eliminate from the target products. Moreover, in many cases, the organocatalysed reactions can be performed in benign solvents and do not require anhydrous conditions. It is well-known that most of the above-mentioned reactions are promoted by a simple aminoacid, l-proline, or, to a lesser extent, by the more complex cinchona alkaloids. However, during the past three decades, other enantiopure natural compounds, the carbohydrates, have been employed as organocatalysts. In the present exhaustive review, the detailed preparation of all the sugar-based organocatalysts as well as their catalytic properties are described.

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By varying the steric and electronic surroundings of the hydrogen-bonding motif, the novel chiral Cinchona-alkaloid based selenoureas were developed. Acting as bifunctional catalysts, they were applied in the Michael reactions of dithiomalonate and nitrostyrene providing chiral adducts with up to 96% ee. The asymmetric Michael--hemiacetalization reaction of benzylidene pyruvate and dimedone, performed with the assistance of 5 mol% of selenoureas, furnished the product with up to 93% ee and excellent yields. The effectiveness of the new hydrogen-bond donors was also proved in solvent-free reactions under ball mill conditions, supporting the sustainability of the devised catalytic protocol.

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ß-thalassemias are among the most common inherited hemoglobinopathies worldwide and are the result of autosomal mutations in the gene encoding ß-globin, causing an absence or low-level production of adult hemoglobin (HbA). Induction of fetal hemoglobin (HbF) is considered to be of key importance for the development of therapeutic protocols for ß-thalassemia and novel HbF inducers need to be proposed for pre-clinical development. The main purpose on this study was to analyze Cinchona alkaloids (cinchonidine, quinidine and cinchonine) as natural HbF-inducing agents in human erythroid cells. The analytical methods employed were Reverse Transcription quantitative real-time PCR (RT-qPCR) (for quantification of γ-globin mRNA) and High Performance Liquid Chromatography (HPLC) (for analysis of the hemoglobin pattern). After an initial analysis using the K562 cell line as an experimental model system, showing induction of hemoglobin and γ-globin mRNA, we verified whether the two more active compounds, cinchonidine and quinidine, were able to induce HbF in erythroid progenitor cells isolated from ß-thalassemia patients. The data obtained demonstrate that cinchonidine and quinidine are potent inducers of γ-globin mRNA and HbF in erythroid progenitor cells isolated from nine ß-thalassemia patients. In addition, both compounds were found to synergize with the HbF inducer sirolimus for maximal production of HbF. The data obtained strongly indicate that these compounds deserve consideration in the development of pre-clinical approaches for therapeutic protocols of ß-thalassemia.

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N-Protected oxindole derivatives of unprecedented malleability bearing ester moieties at C-3 have been shown to participate in enantioselective phase-transfer-catalysed alkylations promoted by ad-hoc designed quaternary ammonium salts derived from quinine bearing hydrogen-bond donating substituents. For the first time in such phase-transfer-catalysed enolate alkylations, the reactions were carried out under base-free conditions. It was found that urea-based catalysts outperformed squaramide derivatives, and that the installation of a chlorine atom adjacent to the catalyst's quinoline moiety aided in avoiding selectivity-reducing complications related to the production of HBr in these processes. The influence of steric and electronic factors from both the perspective of the nucleophile and electrophile were investigated and levels of enantiocontrol up to 90% ee obtained. The synthetic utility of the methodology was demonstrated via the concise enantioselective synthesis of a potent CRTH2 receptor antagonist.

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The substance class of the well-known Cinchona alkaloids is widened by 6'-Amino-cinchonine and 6'-Amino-cinchonidine, novel compounds which incorporate a primary amino function in the quinolinic ring system. These key intermediates open the field for a range of fruitful chemistry. Here is described a short and direct pathway for the synthesis of triazole containing derivatives of the above-mentioned substances using the [3 + 2] Huisgen cycloaddition. For this purpose, the amines were first converted into the corresponding azides. Based on this, non-substituted and silyl-protected triazoles were synthesized as examples. Furthermore, didehydrated derivatives of quincorine and quincoridine were used as addition partners, resulting in compounds that carry the quinuclidine ring of the cinchona alkaloids at both ends. Some of these compounds were examined radiographically to investigate the position of the quinuclidine ring to the triazole. The solid-state structures of compounds 10, 11 and 28 were determined by X-ray diffraction analyses.

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Cinchona alkaloid-derived organocatalysts are widely employed in various asymmetric transformations, yielding products with high enantiopurity. In this respect, a bifunctional quinine-derived sulfonamide organocatalyst was developed to catalyze the asymmetric sulfa-Michael reaction of naphthalene-1-thiol with trans-chalcone derivatives. The target sulfa-Michael adducts were obtained with up to 96% ee under mild conditions and with a low (1 mol %) catalyst loading. Selected enantiomerically enriched sulfa-Michael addition products were subjected to oxidation to obtain the corresponding sulfones.

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We herein report an unprecedented strategy for the asymmetric α-chlorination of ß-keto esters with hypervalent iodine-based Cl-transfer reagents using simple Cinchona alkaloid catalysts. Our investigations support an α-chlorination mechanism where the Cinchona species serves as a nucleophilic catalyst by reacting with the chlorinating agent to generate a chiral electrophilic Cl-transfer reagent in situ. Using at least 20 mol-% of the alkaloid catalyst allows for good yields and enantioselectivities for a variety of different ß-keto esters under operationally simple conditions.

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An efficient approach to the synthesis of chiral selenoureas consisting of Cinchona alkaloid scaffolds was described. The new selenoureas were assessed as bifunctional organocatalysts in the asymmetric Michael addition reactions under mild conditions. The best results were obtained for selenoureas bearing the 4-fluorophenyl group. These catalysts promoted the reactions with enantioselectivities of up to 96% ee. Additionally, the catalytic performance of the thiourea and selenourea counterpart was compared.

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The ability of quinoline alkaloids (cinchonine, cinchonidine, quinine, and quinidine) to sensitize different human cancer cell lines to doxorubicin (DOX)-induced cell death was evaluated. Cell viability was analyzed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the alkaloids ability to enhance DOX-induced apoptosis was explored using Western blotting analysis. Also, flow cytometry was applied to analyze cell fractions in the different cell cycle phases. All alkaloids showed a significant enhancement of DOX-induced cell death in HeLa and HepG2 cell lines. The chemosensitizing activity of the quinoline alkaloids was attributed to the induction of apoptosis as indicated by splitting of caspase-3 and its substrate poly (ADP-ribose) polymerase (PARP). In addition, there was an increase in the cell fractions in sub-G0/G1 phase in case of DOX combination with the alkaloids. This study proves the ability of the quinoline alkaloids to enhance DOX-induced apoptotic cell death in human cervical and hepatocellular carcinoma cells.

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In this study, we present results obtained on the diastereo- and enantioseparation of some basic natural and synthetic Cinchona alkaloid analogs by applying liquid chromatographic (LC) and subcritical fluid chromatographic (SFC) modalities on amylose and cellulose tris-(phenylcarbamate)-based stationary phases using n-hexane/alcohol/DEA or CO2/alcohol/DEA mobile phase systems. Seven chiral stationary phases in their immobilized form were employed to explore their stereoselectivity for a series of closely related group of analytes. The most important characteristics of LC and SFC systems were evaluated through the variation of the applied chromatographic conditions (e.g., the nature and content of the alcohol modifier, the concentration of additives, temperature). The columns Chiralpak IC and IG turned out to be the best in both LC and SFC modalities. Temperature-dependence study indicated enthalpy-controlled separation in most cases; however, separation controlled by entropy was also registered.

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The use of pseudo enantiomers is a well-known method of achieving products of complementary stereochemistry. Only rarely can different enantiomers of a product be accessed without modulation of the catalyst. Recently, a system was reported wherein two different enantiomers of spirocycles were obtained by a cascade reaction of unsaturated pyrrolin-4-ones with mercaptoacetaldehyde catalyzed by a single optimized cinchona alkaloid squaramide-derived organocatalyst. It was originally proposed that the E/Z geometry of the unsaturated pyrrolin-4-one dictated the stereochemistry of the spirocycle product, but this was not investigated further. In the present work, we have investigated the nature of a pseudo-enantiomeric organocatalyst conformation applying density functional theory calculations for investigating the transition states for the reaction. Furthermore, the influence of the double-bond geometry of the pyrrolin-4-one has been studied beyond what is possible to test experimentally. The results provide a greater understanding for this class of reactions that may be applicable in future methodology development.

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In this report, we introduce a new strategy for controlling the stereochemistry in Ugi adducts. Instead of controlling stereochemistry directly during the Ugi reaction we have attempted to stereodefine the chiral center at the peptidyl position through the post-Ugi functionalization. In order to achieve this, we chose to study 2-oxo-aldehyde-derived Ugi adducts many of which partially or fully exist in the enol form that lacks the aforementioned chiral center. This in turn led to their increased nucleophilicity as compared to the standard Ugi adducts. As such, the stereocenter at the peptidyl position could be installed and stereodefined through the reaction with a suitable electrophile. Towards this end, we were able to deploy an asymmetric cinchona alkaloid-promoted electrophilic fluorination producing enantioenriched post-Ugi adducts fluorinated at the peptidyl position.

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α-Aminophosphonic acids are phosphorus analogues of α-amino acids. Compounds of this type find numerous applications in medicine and crop protection due to their unique biological activities. A crucial factor in these activities is the configuration of the stereoisomers. Only a few methods of stereoselective transformation of α-amino acids into their phosphorus analogues are known so far and all of them are based on asymmetric induction, thus involving the use of a chiral substrate. In contrast, we have focused our efforts on the development of an effective method for this type of transformation using a racemic mixture of starting N-protected α-amino acids and a chiral catalyst. Herein, a simple and efficient stereoselective organocatalytic α-amidoalkylation of dimethyl phosphite by 1-(N-acylamino)alkyltriphenylphosphonium salts to enantiomerically enriched α-aminophosphonates is reported. Using 5 mol% of chiral quinine- or hydroquinine-derived quaternary ammonium salts provides final products in very good yields up to 98% and with up to 92% ee. The starting phosphonium salts were easily obtained from α-amino acid derivatives by decarboxylative methoxylation and subsequent substitution with triphenylphosphonium tetrafluoroborate. The appropriate self-disproportionation of enantiomers (SDE) test for selected α-aminophosphonate derivatives via achiral flash chromatography was performed to confirm the reliability of the enantioselectivity results that were obtained.

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Liquid chromatographic (LC) and subcritical fluid chromatographic (SFC) resolution of the basic natural and synthetic Cinchona alkaloid analogues has been studied. Focus has been placed on the employment of four enantiomerically structured chiral strong cation-exchangers and four chiral diastereoisomeric Cinchona alkaloid and cyclohexyl aminosulfonic acid-based zwitterionic ion-exchangers. Except for the novel, recently synthesized racemic quinine the other investigated pairs of basic analytes are diastereomeric, but often called "pseudoenantiomeric" compounds of quinine and quinidine, cinchonidine and cinchonine, 9­epi­quinine and 9­epi­quinidine. As expected, the elution order of the resolved racemic quinine was reversed for all the eight investigated enantiomeric and (pseudo)enantiomeric pairs of chiral stationary phases, whereas this was not necessarily the case for the diastereomeric pairs of the Cinchona alkaloid related analytes. Varying the type and composition of the protic (methanol) and non-protic (acetonitrile) but polar bulk solvents in combination with organic salt additives in the mobile phase the overall retention and stereoselectivity characteristics could be triggered, leading to well performing LC and SFC systems. Thus the retention behavior of the basic analytes on both the chiral cation-exchangers and the diastereomeric zwitterionic ion-exchangers, used as cation-exchangers, could be described by the stoichiometric displacement model related to the counter-ion effect of the mobile phase additives. In addition, it became obvious that the non-protic acetonitrile compared to methanol as bulk solvent lead to a significant increase in retention, which can be associated with an increased electrostatic interaction of the charged sites due to a smaller solvation shell of the solvated cationic and anionic species. Based on the chromatographic results of the systematically selected chiral analytes and stationary phases attempts were undertaken to interpret qualitatively the observed stereoselectivity phenomena.

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The current chemotherapy of Chagas disease needs to be urgently improved. With this aim, a series of 16 hybrids of Cinchona alkaloids and bile acids were prepared by functionalization at position C-2 of the quinoline nucleus by a radical attack of a norcholane substituent via a Barton-Zard decarboxylation reaction. The antitrypanosomal activity of the hybrids was tested on different stages and strains of T. cruzi. In particular, eight out of 16 hybrids presented an IC50 ≤1 µg/mL against trypomastigotes of the CL Brener strain and/or a selectivity index higher than 10. These promising hybrids yielded similar results when tested on trypomastigotes from the RA strain of T. cruzi (discrete typing unit-DTU-VI). Surprisingly, trypomastigotes of the Y strain (DTU II) were more resistant to benznidazole and to most of the hybrids than those of the CL Brener and RA strains. However, the peracetylated and non-acetylated forms of the cinchonine/chenodeoxycholic bile acid conjugate 4f and 5f were the most trypanocidal hybrids against Y strain trypomastigotes, with IC50 values of 0.5 and 0.65 µg/mL, respectively. More importantly, promising results were observed in invasion assays using the Y strain, where hybrids 5f and 4f induced a significant reduction in intracellular amastigotes and on the release of trypomastigotes from infected cells.

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Decarboxylative aldol reaction of aliphatic carboxylic acids is a useful method for C-C bond formation because carboxylic acids are an easily available class of compounds. In this study, we found that the decarboxylative aldol reaction of tertiary ß-ketocarboxylic acids and trifluoropyruvates proceeded smoothly to yield the corresponding aldol products in high yields and with high diastereoselectivity in the presence of a tertiary amine catalyst. In this reaction, we efficiently constructed a quaternary carbon center and an adjacent trifluoromethylated carbon center. This protocol was also extended to an enantioselective reaction with a chiral amine catalyst, and the desired product was obtained with up to 73% enantioselectivity.

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