RESUMEN
In the field of oligonucleotides drug discovery, phosphorothioate (PS) modification has been recognized as an effective tool to overcome the nuclease digestion, and generates 2n of possible diastereomers, where n equals the number of PS linkages. However, it is also well known that differences in drug efficacy and toxicity are caused by differences in stereochemistry of oligonucleotides. Therefore, the development of a high-resolution analytical method that enables stereo discrimination of oligonucleotides is desired. Under this circumstance, capillary electrophoresis (CE) using polyvinylpyrrolidone (PVP) is considered as one of the useful tools for the separation analysis of diastereomers. In this study, we evaluated the several oligonucleotides with the structural diversities in order to understand the separation mechanism of the diastereomers by CE. Especially, five kinds of 2'-moieties were deeply examined by CE with PVP 1,300,000 polymer solution. We found that different trend of the peak shapes and the peak resolution were observed among these oligonucleotides. For example, the better peak resolution was observed in 6 mer PS3-DNA compared to the rigid structure of 6 mer PS3-LNA. As for this reason, the computational simulation revealed that difference of accessible surface area caused by the steric structure of thiophosphate in each oligonucleotide is one of the key attributes to explain the separation of the diastereomers. In addition, we achieved the separation of sixteen peak tops of the diastereomers in 6 mer PS4-DNA, and the complete separation of fifteen diastereomers in 6 mer PS4-RNA. These knowledge for the separation of the diastereomers by CE will be expected to the quality control of the oligonucleotide drugs.
Asunto(s)
Electroforesis Capilar , Oligonucleótidos , Povidona , Electroforesis Capilar/métodos , Estereoisomerismo , Povidona/química , Oligonucleótidos/química , Oligonucleótidos/análisis , Oligonucleótidos/aislamiento & purificaciónRESUMEN
The absolute configuration has been established of the enantiopure anti-head-to-head cyclodimer of anthracene-2-carboxylic acid (AC) cocrystallized with L-propinol and dichloromethane [systematic name: (S)-2-(hydroxymethyl)pyrrolidin-1-ium (5R,6S,11R,12S)-8-carboxy-5,6,11,12-tetrahydro-5,12:6,11-bis([1,2]benzeno)dibenzo[a,e][8]annulene-2-carboxylate dichloromethane monosolvate], C5H12NO(+)·C30H19O4(-)·CH2Cl2. In the crystal structure, the AC dimer interacts with L-prolinol through a nine-membered hydrogen-bonded ring [R2(2)(9)], while the dichloromethane molecule is incorporated to fill the void space. The absolute configuration determined in this study verifies a recent assignment made by comparing theoretical versus experimental circular dichroism spectra.
RESUMEN
Competitive cross-/homo-photocyclodimerization of anthracene (AN) and 2-anthracenecarboxylic acid (AC) mediated by a chiral hydrogen-bonding template (TKS) was investigated under various conditions. The cross-photocyclodimerization was favored by a factor of 4-5 at all temperatures and wavelengths examined to afford the AC-AN cross-dimer in 80-84% yield even at AN/AC = 1 and in 98% yield at AN/AC = 10. The enantiomeric excesses (ee's) obtained were 27-47% for the homo-dimers and 21-24% for the cross-dimer. The absolute configuration of the cross-dimer was determined by comparing the experimental and theoretical circular dichroism spectra and further correlated with the re/si enantiotopic-face selectivity upon AC-TKS complexation in the ground state. Detailed analyses of the complexation behavior and the fluorescence lifetime and cyclodimerization rate of excited re/si complexes revealed that the product's ee is critically controlled not only by the relative abundance of the re/si complexes in the ground and excited states but also by their relative photocyclodimerization rate. Crucially, the ground-state thermodynamics and the excited-state kinetics are not synergistic but offsetting in enantiotopic-face selectivity, and the latter overwhelms the former to give the homo- and cross-dimers in modest ee's. Finally, some practical strategies for enhancing the enantioselectivity in chiral template-mediated photochirogenesis have been proposed.
Asunto(s)
Antracenos/química , Ácidos Carboxílicos/química , Ciclización , Dimerización , Enlace de Hidrógeno , Procesos Fotoquímicos , Teoría Cuántica , Estereoisomerismo , TermodinámicaRESUMEN
Circular dichroism (CD) spectral examinations at various host/guest ratios revealed that 2-anthracenecarboxylic acid (AC) forms not only 1:1 but also novel 2:1 hydrogen-bonded/π-stacked complexes with a chiral 4-benzamidoprolinol template (TKS159). The 2:1 complexation is a minor process but causes significant CD spectral changes as a consequence of the exciton coupling interaction of two AC chromophores and greatly accelerates the head-to-head photocyclodimerization to significantly affect the stereochemical outcomes.
Asunto(s)
Antracenos/química , Ácidos Carboxílicos/química , Pirrolidinas/química , Dicroismo Circular , Dimerización , Enlace de Hidrógeno , Estructura Molecular , Procesos Fotoquímicos , EstereoisomerismoRESUMEN
The photochirogenesis of 2-anthracenecarboxylic acid (AC) complexed to a hydrogen-bonding template (TKS159) was investigated to obtain mechanistic information on how chirogenesis is achieved for the dimerization of AC. Complexation of AC to TKS159 leads to the shielding of one of the two surfaces of the prochiral AC molecule. The two diastereomeric AC-TKS complexes, i.e., re-AC-TKS and si-AC-TKS, were characterized by changes in the UV-vis, fluorescence, and circular dichroism spectra and excited-state lifetimes. The ee is not simply determined by the diastereomeric ratio of the re- and si-AC-TKS complexes but also depends on the relative lifetimes of the diastereomeric complexes. The relative population of the re and si complexes was calculated from the enantiomeric excess (ee) for the products, taking into account the relative lifetimes of the two complexes. These studies established a protocol that can be used to reveal the mechanism for photochirogenesis by investigating the ground state and the excited state behavior of supramolecular systems.
Asunto(s)
Aminas/química , Antracenos/química , Ácidos Carboxílicos/química , Pirrolidinas/química , Ciclización , Dimerización , Enlace de Hidrógeno , Estructura Molecular , Fotoquímica , EstereoisomerismoRESUMEN
The interaction of a newly developed Helicobacter pylori eradicating agent (TG44, 4-methylbenzyl-4'-[trans-4-(guanidinomethyl)cyclohexylcarbonyloxy]biphenyl-4-carboxlylate monohydrochloride) with beta-cyclodextrin (beta-CyD) in aqueous solution and in solid state was studied to gain insight into the high in-vivo H. pylori eradicating activity of TG44/beta-CyD complex. The interaction was studied by the solubility method, spectroscopic methods, powder X-ray diffractometry and differential scanning colorimetry (DSC). TG44 gave A(L)-type phase solubility diagram with beta-CyD in water, showing a linear increase in solubility of the drug up to 8 mM beta-CyD concentration. The solubility of TG44 (0.04 mM in water at 25 degrees C) increased about 70-folds at 8 mM beta-CyD. Ultraviolet, circular dichroism, fluorescence and (1)H-nuclear magnetic resonance spectroscopic studies indicated that TG44 forms the inclusion complex with beta-CyD in a 1:1 stoichiometry and the biphenyl moiety of TG44 is preferably included in the beta-CyD cavity in water. The Giordano plot made by monitoring changes in the fusion enthalpy of TG44 (about 184 degrees C) suggested that TG44 forms the 1:1 complex with beta-CyD in the solid state. The TG44/beta-CyD solid complex in a 1:1 stoichiometry was prepared by the grinding and spray-drying methods and confirmed by powder X-ray diffractometry and DSC that the complex is in an amorphous state. The initial dissolution rate of TG44/beta-CyD complex was significantly faster than those of the drug alone and the physical mixture of both components, maintaining higher supersaturated concentrations of the drug for a long time. The results suggested that the higher eradicating activity of TG44/beta-CyD complex to Helicobacter pylori, compared with that of the drug alone, is attributable at least partly to the faster dissolving property of the complex and its ability to maintain the supersaturated state of the drug in the gastric fluid.
Asunto(s)
Benzoatos/química , Guanidinas/química , Agua/química , beta-Ciclodextrinas/química , Benzoatos/farmacología , Rastreo Diferencial de Calorimetría , Dicroismo Circular , Guanidinas/farmacología , Helicobacter pylori/efectos de los fármacos , Resonancia Magnética Nuclear Biomolecular , Solubilidad , Temperatura , Factores de Tiempo , Difracción de Rayos X , beta-Ciclodextrinas/farmacologíaRESUMEN
[Structure: see text] Supramolecular enantiodifferentiating photocyclodimerization of 2-anthracenecarboxylic acid (AC) was performed in the presence of (2S,4S)-4-amino-5-chloro-2-methoxy-N-(1-ethyl-2-hydroxymethyl-4-pyrrolidinyl)benzamide (TKS159), and its stereoisomers were employed as chiral templates. The TKS template provides us with a novel hydrogen-bonding and shielding motif for enantioface-selectively binding an AC molecule. Chiral products 2 and 3 were obtained in good enantiomeric excesses (ee's) of 40% and 40%, respectively.
RESUMEN
Due to concerns about the current therapeutic modalities for Helicobacter pylori infection, e.g., the increased emergence of drug-resistant strains and the adverse reactions of drugs currently administered, there is a need to develop an anti-H. pylori agent with higher efficacy and less toxicity. The antibacterial activity of TG44, an anti-H. pylori agent with a novel structural formula, against 54 clinical isolates of H. pylori was examined and compared with those of amoxicillin (AMX), clarithromycin (CLR), and metronidazole (MNZ). Consequently, TG44 inhibited the growth of H. pylori in an MIC range of 0.0625 to 1 microg/ml. The MIC ranges of AMX, CLR, and MNZ were 0.0078 to 8 microg/ml, 0.0156 to 64 microg/ml, and 2 to 128 microg/ml, respectively. The antibacterial activity of TG44 against AMX-, CLR-, and MNZ-resistant strains was nearly comparable to that against drug-susceptible ones. In a pH range of 3 to 7, TG44 at 3.13 to 12.5 microg/ml exhibited potent bactericidal activity against H. pylori in the stationary phase of growth as early as 1 h after treatment began, in contrast to AMX, which showed no bactericidal activity at concentrations of up to 50 microg/ml at the same time point of treatment. TG44 at 25 microg/ml exhibited no antibacterial activity against 13 strains of aerobic bacteria, suggesting that its antibacterial activity against H. pylori is potent and highly specific. The present study indicated that TG44 possesses antibacterial activity which manifests quickly and is potentially useful for eradicating not only the antibiotic-susceptible but also the antibiotic-resistant strains of H. pylori by monotherapy.
Asunto(s)
Benzoatos/farmacología , Guanidinas/farmacología , Helicobacter pylori/efectos de los fármacos , Amoxicilina/farmacología , Claritromicina/farmacología , Farmacorresistencia Bacteriana , Helicobacter pylori/aislamiento & purificación , Helicobacter pylori/ultraestructura , Humanos , Concentración de Iones de Hidrógeno , Metronidazol/farmacología , Pruebas de Sensibilidad Microbiana , Microscopía Electrónica de Rastreo , Estructura MolecularRESUMEN
The interaction of a newly developed Helicobacter pylori eradicating agent (TG44, 4-methylbenzyl-4'-[trans-4-(guanidinomethyl)cyclohexylcarbonyloxy]-biphenyl-4-carboxlylate monohydrochloride) with cyclomaltoheptaose (beta-cyclodextrin, beta-CyD) in the solid state was studied by high-speed frequency-switched Lee-Goldburg (FSLG) (13)C-(1)H heteronuclear correlation (HETCOR) NMR experiments. The TG44/beta-CyD solid complex in a 1:1 stoichiometry was prepared by the grinding method. Powder X-ray diffractometry confirmed that the complex is in an amorphous state. The solid-state (13)C signals of TG44 and beta-CyD were significantly broadened by the complexation. As the temperature increased, the (13)C signals of the aromatic moieties of TG44 were insignificantly influenced, whereas those of the cyclohexyl moiety became sharper. The T1(rho) H values of the aromatic moieties of TG44 were almost the same as those of the beta-CyD carbons, whereas those of other TG44 carbons gave much smaller values. The (13)C-(1)H HETCOR spectra gave the intermolecular correlation peaks between the aromatic carbons of TG44 and the beta-CyD protons or between the biphenyl protons of TG44 and the beta-CyD carbons, when measured using longer contact times (500 and 1500mus). On the basis of these solid NMR spectroscopic data together with aqueous NMR data, we assume that beta-CyD includes predominantly the biphenyl moiety of TG44 in the solid state. (13)C-(1)H HETCOR spectroscopy is particularly useful for the determination of inclusion modes of the complexes that occurring in an amorphous form.