Ionic Partition Diagram of the Zwitterionic Antihistamine Cetirizine

A recent analysis of the lipophilicity profile of cetirizine in the octanol/water and dodecane/water systems revealed a partial intramolecular charge neutralization that can partly explain why cetirizine has pharmacokinetic properties differing from those of first‐generation antihistamines such as hydroxyzine. As conformational changes are the principal driving force for this intramolecular effect, the present study deals with the partitioning of cetirizine and hydroxyzine in an apolar medium well‐suited to reveal intramolecular interactions, namely the 1,2‐dichloroethane/water system. The lipophilicity of the different electrical forms of cetirizine and hydroxyzine was studied by two‐phase titrimetry and cyclic voltammetry. The differences in lipophilicity between the dicationic, monocationic, zwitterionic, and anionic species of cetirizine indicated intramolecular interactions via folded conformations, which render the molecule markedly more lipophilic than expected at physiological pH. Folded conformations were also found to predominate in monocationic and neutral hydroxyzine. The ionic partition diagram of cetirizine indicates that it acts as a proton transporter across interfaces under certain conditions of pH and Galvani potential difference. This study underlines the importance of conformational effects on the partition properties of cetirizine and hydroxyzine, as well as the complexity of its interfacial mechanisms of transfer. In particular, cetirizine can facilitate proton transfer, a property of potential biological relevance.     

A fully validated method for the simultaneous determination of 11 antihistamines in breast milk by gas chromatography–mass spectrometry

Antihistamines are excreted into breast milk in small amounts; however, there are no adequate published studies or data concerning their effects on newborns and safety during breastfeeding. Thus, the development of sensitive and specific methodologies for the determination of antihistamines in breast milk is critical. A simple and sensitive GC–MS method for the simultaneous determination of 11 antihistamines (diphenhydramine, orphenadrine, chlorpheniramine, dimethindene, meclozine, hydroxyzine, loratadine, desloratadine, cetirizine, rupatadine and ebastine) in breast milk was developed and validated. The antihistamines were solid‐phase extracted and derivatized with acetic anhydride and n‐propanol. Diazepam‐d₅, hydroxyzine‐d₄ and cetirizine‐d₈ were used as internal standards. Absolute recovery values for all analytes ranged from 70.5 to 120.0%, while the limits of detection and quantification for all analytes were 1.50 and 5.00 ng/mL, respectively. All calibration curves were linear (R² ≥ 0.990) within the range 5.00–1000.0 ng/mL. Accuracy (E_r) ranged between −7.6 and 7.0%, while precision (RSD) was <12% for all antihistamines. The developed method is suitable for the investigation of antihistamine‐related clinical cases, as well as for pharmacokinetic and breastfeeding safety studies.     

Synthesis of Zinc Chlorophyll Homo/Hetero‐Dyads and their Folded Conformers with Porphyrin,Chlorin, and Bacteriochlorin π‐Systems

Zinc complex of pyropheophorbide‐b, a derivative of chlorophyll‐b, was covalently dimerized through ethylene glycol diester. The synthetic homo‐dyad was axially ligated with two methanol molecules from the β‐face and both the diastereomerically coordinating methanol species were hydrogen bonded with the keto‐carbonyl groups of the neighboring chlorin in a complex. The resulting folded conformer in a solution was confirmed by visible, ¹H NMR and IR spectra. All the synthetic zinc chlorin homo‐ and hetero‐dyads consisting of pyropheophorbides‐a, b and/or d took the above methanol‐locked and π–π stacked supramolecules in 1% (v/v) methanol and benzene to give redmost (Qy) electronic absorption band(s) at longer wavelengths than those of the corresponding monomeric chlorin composites. The other zinc chlorin and bacteriochlorin homo‐dyads completely formed similar folded conformers in the same solution, while zinc inverse chlorin and porphyrin homo‐dyads partially took such supramolecules. The J‐type aggregation to folded conformers and the redshift values of composite Qy bands were dependent on the electronic and steric factors of porphyrinoid moieties in dyads.     

Targeting a Dark Excited State of HIV‐1 Nucleocapsid by Antiretroviral Thioesters Revealed by NMR Spectroscopy

HIV‐1 nucleocapsid (NCp7) is a two Cys₂HisCys zinc knuckle (N‐Zn and C‐Zn) protein that plays a key role in viral replication. NCp7 conformational dynamics is characterized by NMR relaxation dispersion and chemical exchange saturation transfer measurements. While the N‐Zn knuckle is conformationally stable, the C‐Zn knuckle interconverts on the millisecond timescale between the major state, in which the zinc is coordinated by three cysteines and a histidine, and two folded minor species (with populations around 1 %) in which one of the coordination bonds (Cys413‐Sγ‐Zn or His421‐N?2‐Zn) is hydrolyzed. These findings explain why antiretroviral thioesters specifically disrupt the C‐Zn knuckle by initial acylation of Cys413, and show that transient, sparsely‐populated (“dark”), excited states of proteins can present effective targets for rational drug design.     

Deciphering the Conformational Choreography of Zinc Coordination Complexes with Standard and Novel Proton NMR Techniques Combined with DFT Methods

The presence of water has been shown to deeply impact the stability and geometry of Zn complexes in solution. Evidence for tetra‐ and penta‐coordinated species in a pyridylmethylamine–Zn^II model complex is presented. Novel ¹H NMR tools such as T₁‐filtered selective exchange spectroscopy and pure shifted gradient‐encoded selective refocusing as well as classical 2D (¹H–¹H) exchange spectroscopy, diffusion‐ordered spectroscopy and T₁(¹H) measurements, in combination with density functional theory methods allow the full conformational dynamics of a pyridylmethylamine–Zn^II complex to be revealed. Four conformers and two families of complexes depending on the hydration states are elucidated.     

Mechanochromism,Twisted/Folded Structure Determination,and Derivatization of (N‐Phenylfluorenylidene)acridane

(N‐Phenylfluorenylidene)acridane (Ph‐FA) compounds with electron‐withdrawing and ‐donating substituents (H, MeO, Ph, NO₂, Br, F) at the para position of the phenyl group were successfully synthesized by Barton–Kellogg reactions of N‐aryl thioacridones and diazofluorene. By using the substituent on the nitrogen atom to alter the electronic properties, both the folded and twisted conformers of p‐NO₂‐C₆H₄‐FA could be crystallographically characterized, which enabled the charge transfer from the electron‐donating acridane moiety to the electron‐accepting fluorenylidene moiety to be understood. Ground‐state mechanochromism, thermochromism, vapochromism, and proton‐induced chromism were demonstrated between the folded and twisted conformations of the conformers. Protonation and chemical oxidation of Ph‐FA gave two stable acridinium compounds, namely, the fluorenylacridinium and acridinium radical cations. The present study will contribute to the development of functional dyes and organic semiconductors.     

Mechanochromism,Twisted/Folded Structure Determination,and Derivatization of (N‐Phenylfluorenylidene)acridane

(N‐Phenylfluorenylidene)acridane (Ph‐FA) compounds with electron‐withdrawing and ‐donating substituents (H, MeO, Ph, NO₂, Br, F) at the para position of the phenyl group were successfully synthesized by Barton–Kellogg reactions of N‐aryl thioacridones and diazofluorene. By using the substituent on the nitrogen atom to alter the electronic properties, both the folded and twisted conformers of p‐NO₂‐C₆H₄‐FA could be crystallographically characterized, which enabled the charge transfer from the electron‐donating acridane moiety to the electron‐accepting fluorenylidene moiety to be understood. Ground‐state mechanochromism, thermochromism, vapochromism, and proton‐induced chromism were demonstrated between the folded and twisted conformations of the conformers. Protonation and chemical oxidation of Ph‐FA gave two stable acridinium compounds, namely, the fluorenylacridinium and acridinium radical cations. The present study will contribute to the development of functional dyes and organic semiconductors.     

Uncovering Key Structural Features of an Enantioselective Peptide‐Catalyzed Acylation Utilizing Advanced NMR Techniques

We report on a detailed NMR spectroscopic study of the catalyst‐substrate interaction of a highly enantioselective oligopeptide catalyst that is used for the kinetic resolution of trans‐cycloalkane‐1,2‐diols via monoacylation. The extraordinary selectivity has been rationalized by molecular dynamics as well as density functional theory (DFT) computations. Herein we describe the conformational analysis of the organocatalyst studied by a combination of nuclear Overhauser effect (NOE) and residual dipolar coupling (RDC)‐based methods that resulted in an ensemble of four final conformers. To corroborate the proposed mechanism, we also investigated the catalyst in mixtures with both trans‐cyclohexane‐1,2‐diol enantiomers separately, using advanced NMR methods such as T₁ relaxation time and diffusion‐ordered spectroscopy (DOSY) measurements to probe molecular aggregation. We determined intramolecular distance changes within the catalyst after diol addition from quantitative NOE data. Finally, we developed a pure shift EASY ROESY experiment using PSYCHE homodecoupling to directly observe intermolecular NOE contacts between the trans‐1,2‐diol and the cyclohexyl moiety of the catalyst hidden by spectral overlap in conventional spectra. All experimental NMR data support the results proposed by earlier computations including the proposed key role of dispersion interaction.     

Polymorphs of the antiviral drug ganciclovir

Ganciclovir (GCV; systematic name: 2‐amino‐9‐{[(1,3‐dihydroxypropan‐2‐yl)oxy]methyl}‐6,9‐dihydro‐1H‐purin‐6‐one), C₉H₁₃N₅O₄, an antiviral drug for treating cytomegalovirus infections, has two known polymorphs (Forms I and II), but only the structure of the metastable Form II has been reported [Kawamura & Hirayama (2009). X‐ray Struct. Anal. Online , 25 , 51–52]. We describe a successful preparation of GCV Form I and its crystal structure. GCV is an achiral molecule in the sense that its individual conformers, which are generally chiral objects, undergo fast interconversion in the liquid state and cannot be isolated. In the crystalline state, GCV exists as two inversion‐related conformers in Form I and as a single chiral conformer in Form II. This situation is similar to that observed for glycine, also an achiral molecule, whose α‐polymorph contains two inversion‐related conformers, while the γ‐polymorph contains a single conformer that is chiral. The hydrogen bonds are exclusively intermolecular in Form I, but both inter‐ and intramolecular in Form II, which accounts for the different molecular conformations in the two polymorphs.     

Kinetic Cooperativity in Human Pancreatic Glucokinase Originates from Millisecond Dynamics of the Small Domain

The hallmark of glucokinase (GCK), which catalyzes the phosphorylation of glucose during glycolysis, is its kinetic cooperativity, whose understanding at atomic detail has remained open since its discovery over 40 years ago. Herein, by using kinetic CPMG NMR spectroscopic data for 17 isoleucine side chains distributed over all parts of GCK, we show that the origin of kinetic cooperativity is rooted in intramolecular protein dynamics. Residues of glucose‐free GCK located in the small domain displayed distinct exchange behavior involving multiple conformers that are substantially populated (p>17 %) with a k_ex value of 509±51 s^?1, whereas in the glucose‐bound form these exchange processes were quenched. This exchange behavior directly competes with the enzymatic turnover rate at physiological glucose concentrations, thereby generating the sigmoidal rate dependence that defines kinetic cooperativity.     

Conformational studies of dammarane‐type triterpenoids using computational and NMR spectroscopic methods

Natural triterpenoids are of great interest to researchers of various fields as they possess diverse physicochemical and biological properties. In medicinal chemistry, detailed information about the chemical structures of bioactive triterpenoids often helps find new lead compounds. Herein, the low‐energy structures of (20S)‐protopanaxadiol and (20S)‐protopanaxatriol, the aglycones of various triterpenoid saponins found in Panax ginseng, and their (20R)‐epimers have been predicted by the geometry optimization of the conformers extracted from molecular dynamics simulations with the self‐consistent‐charge density functional tight‐binding method. By performing quantum mechanical calculations on the low‐energy conformers, we have estimated the NMR chemical shifts of the compounds, which display good agreement with the most recently reported experimental values within an expected range of errors. Our results indicate that theoretical estimation of the NMR parameters of a relatively large molecule with a molecular mass of 500 is feasible. Copyright © 2015 John Wiley & Sons, Ltd.     

Two‐Dimensional Infrared Spectroscopy Reveals the Structure of an Evans Auxiliary Derivative and Its SnCl4 Lewis Acid Complex

Determining the structure of reactive intermediates is the key to understanding reaction mechanisms. To access these structures, a method combining structural sensitivity and high time resolution is required. Here ultrafast polarization‐dependent two‐dimensional infrared (P2D‐IR) spectroscopy is shown to be an excellent complement to commonly used methods such as one‐dimensional IR and multidimensional NMR spectroscopy for investigating intermediates. P2D‐IR spectroscopy allows structure determination by measuring the angles between vibrational transition dipole moments. The high time resolution makes P2D‐IR spectroscopy an attractive method for structure determination in the presence of fast exchange and for short‐lived intermediates. The ubiquity of vibrations in molecules ensures broad applicability of the method, particularly in cases in which NMR spectroscopy is challenging due to a low density of active nuclei. Here we illustrate the strengths of P2D‐IR by determining the conformation of a Diels–Alder dienophile that carries the Evans auxiliary and its conformational change induced by the complexation with the Lewis acid SnCl₄, which is a catalyst for stereoselective Diels–Alder reactions. We show that P2D‐IR in combination with DFT computations can discriminate between the various conformers of the free dienophile N‐crotonyloxazolidinone that have been debated before, proving antiperiplanar orientation of the carbonyl groups and s‐cis conformation of the crotonyl moiety. P2D‐IR unequivocally identifies the coordination and conformation in the catalyst–substrate complex with SnCl₄, even in the presence of exchange that is fast on the NMR time scale. It resolves a chelate with the carbonyl orientation flipped to synperiplanar and s‐cis crotonyl configuration as the main species. This work sets the stage for future studies of other catalyst–substrate complexes and intermediates using a combination of P2D‐IR spectroscopy and DFT computations.     

A Fullerene‐Based Molecular Torsion Balance for Investigating Noncovalent Interactions at the C60 Surface

To investigate the nature and strength of noncovalent interactions at the fullerene surface, molecular torsion balances consisting of C₆₀ and organic moieties connected through a biphenyl linkage were synthesized. NMR and computational studies show that the unimolecular system remains in equilibrium between well‐defined folded and unfolded conformers owing to restricted rotation around the biphenyl C?C bond. The energy differences between the two conformers depend on the substituents and is ascribed to differences in the intramolecular noncovalent interactions between the organic moieties and the fullerene surface. Fullerenes favor interacting with the π‐faces of benzenes bearing electron‐donating substituents. The correlation between the folding free energies and corresponding Hammett constants of the substituents in the arene‐containing torsion balances reflects the contributions of the electrostatic interactions and dispersion force to face‐to‐face arene–fullerene interactions.     

Synthesis of Ethylene‐bridged Heterocyclic Bidentate P(III)N‐Ligands — Oxidation and Complexation Studies

A series of phosphor(III)inanone ligands 4‐7 , linked by ethylene bridges between the nitrogen atoms of the heterocyclic rings, were synthesized by the reaction of the bis‐PCl derivative 3 with the appropriate trimethylsilylamines. The bis‐phosphor(V)inanone compounds 8‐11 were obtained by the oxidation of 4‐7 with hexafluoroacetone (HFA). Oxidation of 4 and 6 with tetrachloro‐orthobenzoquinone (TOB) gave the bis‐phosphor(V)inanones 12 and 13 . The reaction of 4‐6 with [Pt(COD)Cl₂] led to the platinum complexes 14‐16 . All the σ³‐phosphorinanone compounds 4‐7 and the σ⁵‐phosphorinanone compounds 8‐10 , 12 and 13 exist as a mixture of two conformers, as indicated by two signals in the ³¹P‐NMR spectra. However, compounds 9 and 11 exist as single conformers, both display only one sharp singlet in the ³¹P‐NMR spectra. The Pt‐complexes 15 and 16 contain two conformers; one conformer of 16 could be isolated by crystallization. X‐ray crystal structure determinations for compounds 8 , 14 and 16 were conducted, revealing inversion symmetry for 8 and cis arrangement for 14 and 16 .     

NMR‐based conformational analysis of perezone and analogues

Complete assignment of the ¹H NMR chemical shift and coupling constant values of perezone (1), O‐methylperezone (2) and 6‐hydroxyperezone (3) was carried out by total‐line‐shape‐fitting calculations using the PERCH iterative spectra analysis software (PERCH Solutions Ltd., Kuopio, Finland). The resulting simulated spectra for the three compounds showed strong similarity to their corresponding experimental spectra. Particularly, all vicinal, allylic and homoallylic coupling constant values for the side chain of the three compounds were very similar, thus revealing that the conformation of these three molecules in solution is indeed almost identical. This fact is in agreement with extended side chain conformations over folded chain conformations because 1, 2 and 3 undergo completely different intramolecular cycloaddition reactions. In addition, results of double pulsed field gradient spin echo NOESY 1D experiments performed on perezone (1) were unable to provide evidence for folded conformers. Copyright © 2013 John Wiley & Sons, Ltd.     

Structural Insights into the Trp‐Cage Folding Intermediate Formation

The 20 residue long Trp‐cage is the smallest protein known, and thus has been the subject of several in vitro and in silico folding studies. Here, we report the multistate folding scenario of the miniprotein in atomic detail. We detected and characterized different intermediate states by temperature dependent NMR measurements of the ¹⁵N and ¹³C/¹⁵N labeled protein, both at neutral and acidic pH values. We developed a deconvolution technique to characterize the invisible—fully folded, unfolded and intermediate—fast exchanging states. Using nonlinear fitting methods we can obtain both the thermodynamic parameters (ΔH^F–I, T_m^F–I, ΔC_p^F–I and ΔH^I–U, T_m^I–U, ΔC_p^I–U) and the NMR chemical shifts of the conformers of the multistate unfolding process. During the unfolding of Trp‐cage distinct intermediates evolve: a fast‐exchanging intermediate is present under neutral conditions, whereas a slow‐exchanging intermediate‐pair emerges at acidic pH. The fast‐exchanging intermediate has a native‐like structure with a short α‐helix in the G¹¹–G¹⁵ segment, whereas the slow‐exchanging intermediate‐pair presents elevated dynamics, with no detectable native‐like residue contacts in which the G¹¹? P¹² peptide bond has either cis or trans conformation. Heteronuclear relaxation studies combined with MD simulations revealed the source of backbone mobility and the nature of structural rearrangements during these transitions. The ability to detect structural and dynamic information about folding intermediates in vitro provides an excellent opportunity to gain new insights into the energetic aspects of the energy landscape of protein folding. Our new experimental data offer exceptional testing ground for further computational simulations.     

NMR structural characterization from one‐bond 13C?13C couplings: Complete assignment of a hydrogen‐poor depsidone

The connectivity, conformation, tautomeric form, and dynamics of a new depsidone (perisalazinic acid) were characterized using one‐bond ¹³C? ¹³C NMR scalar couplings (¹J_CC) obtained from the INADEQUATE experiment. Characterization of perisalazinic acid using more conventional NMR techniques is problematic due to the extremely limited number of C? H protons present. In the present study, 81 candidate structures were considered and a best fit structure was selected by comparing computed ¹J_CC values for each candidate to 15 experimental values. Of the six flexible moieties in perisalazinic acid, three are adequately represented by a single orientation stabilized by intramolecular hydrogen bonding. The three remaining groups are present as mixtures of conformers with two sites consisting of a pair of conformations and another disordered over six orientations. This study demonstrates the feasibility of complete three‐dimensional structural characterization of an unknown using only theoretical and experimental ¹J_CC values.     

Candibirin A,a furano­coumarin dimer isolated from Heracleum candicans Wall.

Candibirin A [systematic name: 9,9′‐(1,4‐dioxane‐2,5‐diyldi­methyl­ene­dioxy)­di(7H‐furo­[3,2‐g]­chromen‐7‐one)], a new furano­coumarin dimer, was isolated from Heracleum candicans Wall . ¹H NMR and MS spectra had indicated that the title compound was a dimer of heraclenin or heraclenol, but the linkage structure and its chirality were undetermined. The dioxane linkage, having the R,R configuration, has now been elucidated from di­methyl sulfoxide‐solvated crystals, C₃₂H₂₈O₁₀·2C₂H₆OS. Candibirin A is thus a dimerization product from heraclenin formed by reaction at the epoxy group. Di­methyl­form­amide‐solvated crystals, C₃₂H₂₈O₁₀·C₃H₇NO, adopt a different conformation, with a folded structure that differs from the extended structure in the dimethyl sulfoxide solvate. However, the puckering of the dioxane linker unit is very similar in the two conformers. This result shows that the rotation of the ether bonds, in the linker between the furano­coumarin and dioxane moieties, causes the conformational flexibility of (I).     

Conformational analysis of some 1R,4S‐2‐arylidene‐p‐menthan‐3‐ones by 1H NMR spectroscopy and molecular simulation

Based on ¹H NMR spectral analysis combined with molecular simulation, conformational states of the cyclohexanone ring were studied for some 1R,4S‐2‐(4‐X‐benzylidene)‐p‐menthan‐3‐ones (X = COOCH₃ or C₆H₅) in CDCl₃ and C₆D₆. The co‐existence of chair conformers with an axial orientation of both alkyl substituents and twist‐boat forms was established for the compounds studied at room temperature (22–23° C). The substituent X does not influence appreciably the ratio of these conformers, but the fraction of twist‐boat forms increases noticeably in benzene solutions as compared with CDCl₃ solutions. Rotameric states of the isopropyl fragment were also characterised for the compounds studied. Distinctions in conformational states for the 1R,4S‐2‐arylidene‐p‐menthan‐3‐ones and (?)‐menthone were revealed and are discussed. Copyright © 2002 John Wiley & Sons, Ltd.     

Single‐Point Remote Control of Flapping Motion in Clothespin‐Shaped Bimetallic Palladium Complexes Based on the N(sp2)–N(sp3) Interconversion in Amide Functionalities

The synthesis, structure, and flapping motion of clothespin‐shaped binuclear trans‐bis(salicylaldiminato)palladium(II) complexes (anti‐ 1 ) with 4‐azaheptamethylene linkers bearing amide ( a – g ), urethane ( h ), or urea ( i ) functionalities are described in this report. Various 2D ¹H NMR experiments and XRD analyses indicate that the amide‐ and urethane‐linked anti‐ 1 a , b , d – h complexes exist as equilibrated mixtures of major and minor conformers I and II in CDCl₃, whereas the complexes anti‐ 1 c and i were observed as a single species. The mapping of NOESY cross‐peaks between conformers I and II revealed that the equilibration of the major and minor conformers of anti‐ 1 a , b , d – h proceeds by two pathways, namely a nonrotatory flapping motion of the coordinated blades and a nonflapping rotation of C?N bonds, whereas the equilibration of anti‐ 1 c proceeds by simultaneous flapping and rotation motions. Kinetic studies carried out by means of ¹H–¹H EXSY experiments revealed that 1) the ΔG^≠_298K values for the flapping motion are controlled remotely by the steric and electronic effects of the RCON functionalities and 2) the activation parameters for the nonrotatory flapping process are identical to those for the nonflapping peptide rotation in the complexes anti‐ 1 a,b,d – h , which indicates that the present multistep conformational transformation induced by the flapping motion is controlled by the rate‐determining pyramidalization/depyramidalization (i.e., sp²/sp³ interconversion) of the nitrogen atoms of the functionalities. The static and controllable molecular mobility of anti‐ 1 bearing peptide linkers has been discussed by comparison with the dynamic behavior of its analogues anti‐ 2 – 4 with flexible polymethylene linkers.