Accuracy and sensitivity of the determination of rate constants of interconversion in achiral and chiral environments by dynamic enantioselective electrophoresis

The method for determination of rate constants of interconversion of enantiomers in chiral and achiral environments of a dynamic enantioseparation system was investigated in order to reveal its accuracy, sensitivity and robustness. Two significantly different enantioseparation systems were selected, one with a single (well-defined) chiral selector (CS) and the second with a mixture of CSs, and the rate constants of interconversion for these two systems were compared statistically. While the rate constants of interconversion in the chiral environment were found to be significantly different, the rate constants in achiral environment were confirmed to be statistically the same. The accuracy of the method was independent of experimental conditions. Influence of a CS and temperature on the rate of interconversion were discussed within the scope of determined thermodynamic parameters and statistical evaluation. A certain temperature may exist at which two different types of CSs influence the rate of interconversion equally while the extent of their influence may largely differ at other temperatures.     

On-flow gas chromatographic method for the determination of the enantiomer interconversion energy barrier

A novel on-flow gas chromatographic (GC) method is developed for the determination of the kinetic rate constants and interconversion energy barrier of thermally labile enantiomers. The validity of the developed method is approved by the study of interconversion of 1-chloro-2,2-dimethylaziridine enantiomers on an achiral column. The overall experiments are performed in a series of three columns placed in two independently heated GC ovens. The racemate of the 1-chloro-2,2-dimethylaziridine is injected and separated in the first chiral column at 60 degrees C in which the interconversion of enantiomers is suppressed. Separated enantiomers are then transferred into the achiral column, where the enantiomers are interconverted at a selected temperature under the current carrier gas flow. Effluent from this column is transferred into the second chiral column, where the native enantiomers and those originated by the on-flow interconversion on an achiral column are again separated at 60 degrees C. Chromatograms obtained by monitoring the effluents from the second chiral column are used to determine the peak areas of the original and the newly interconverted enantiomers. The corresponding peak areas and the interconversion times are used to calculate the interconversion rate constants and energy barriers of the 1-chloro-2,2-dimethylaziridine enantiomers. The apparent energy barriers of the enantiomers of 1-chloro-2,2-dimethylaziridine are equal for both enantiomers within a 95% confidence interval and independent of the polarity of the stationary phase of the column in which the interconversion of enantiomers occur.     

Gas chromatographic determination of the interconversion energy barrier for dimethyl-2,3-pentadienedioate enantiomers

The enantiomers of dimethyl-2,3-pentadienedioate undergo interconversion during gas chromatographic separation on 2,6-di-O-methyl-3-O-pentyl-beta-, 2,6-di-O-methyl-3-O-pentyl-gamma-, and 2,3-di-O-methyl-6-O-tert butyldimethylsilyl-beta-CD chiral stationary phases. The combination of a deconvolution method with an internal standard was used to determine individual enantiomer peak areas and retention times needed for the calculation of the interconversion rate constants and the energy barrier for dimethyl-2,3-pentadienedioate enantiomers. The kinetic and thermodynamic data obtained for the interconversion data (rate constants, energy barriers, enthalpies, and entropies) were in good agreement with the published data (Trapp, O., Schurig, V., Chirality 2002, 14, 465-470) using permethylated-beta-CD (Chirasil-beta-Dex).     

Direct measurement of the rate of interconversion of zirconaaziridine enantiomers

The insertion of enantiopure C2-symmetric diphenylethylene carbonate into the Zr-C bonds of zirconaaziridines leads to the asymmetric synthesis of amino acid methyl esters. Because the zirconaaziridine enantiomers interconvert, the reaction is a dynamic kinetic resolution (DKR). The efficiency of the DKR (the ratio of the two diastereomeric products) is determined by the balance between the rate of enantiomer interconversion and the rate of insertion; slow addition of the inserting enantiopure carbonate is often required to maximize the stereoselectivity. For a case when enantiomer interconversion is fast, its rate constant kinv has been determined by NMR line broadening; for a case when interconversion is slow, k(inv) has been determined by computer simulation of the formation of the diastereomeric products as a function of time; for several intermediate cases, k(inv) has been determined by making the zirconaaziridine enantioenriched and monitoring its racemization by CD spectroscopy. The observed k(inv) is independent of [THF], implying that interconversion occurs with THF coordinated. Interconversion presumably occurs via an achiral intermediate, either a rapidly inverting (via an eta1-N structure) eta3-azaallyl hydride or an eta1-imine. As addition of THF slows insertion without affecting enantiomer interconversion, it produces a more efficient DKR without slow addition of the enantiopure carbonate.     

Determination of the 2,3-pentadienedioic acid enantiomer interconversion energy barrier 1. Classical kinetic approach

A classical kinetic method was used to determine the energy barrier for the inter-conversion of 2,3-pentadienedioic acid enantiomers. Each individual enantiomer was isolated by collecting the appropriate peaks from the HPLC enantiomeric separation, of racemic 2,3-pentadienedioic acid. The isolated enantiomers were racemized at 22 degrees C using various interconversion times. The ratio of enantiomers in each reaction solution was determined by HPLC at 22 degrees C. The corresponding peak areas of the enantiomers and the interconversion times obtained from the HPLC chromatograms were used to calculate both the interconversion rate constants describing (+)--> (-) and (-) --> (+) interconversions as well as the energy barriers. It was confirmed that the interconversion of 2,3-pentadienedioic acid enantiomers is a firstorder kinetic reaction. Both semiempirical and ab initio methods were used to explore the mechanism of the interconversion of 2,3-pentadienedioic acid enantiomers, and to calculate the interconversion energy barrier. Comparison of the interconversion energy barriers found by the ab initio method (deltaG# = 110.7 kJ/mol) and by classical kinetics in the mobile phase solution at 22 degrees C (delta Gapp = 93.9+/-0.2 kJ/mol) shows a difference which may be attributed to the different conditions assumed in the theoretical calculation (i.e., a gaseous state) and the actual experimental conditions (i. e., liquid solution) and a possible catalytic effect of the solution composition.     

Dynamics of interconversion of enantiomers in chiral separation systems: a novel approach for determination of all rate constants involved in the interconversion

When enantiomers separated by chromatography or capillary electrophoresis undergo interconversion reaction (enantiomerization) during the separation, it leads to a typical detection pattern: two individual peaks of the separated enantiomers are connected with a plateau consisting of a mixture of both separated enantiomers. We propose a separation method for determination of all individual rate constants (or inversion barriers) of the interconversion. The method enables to distinguish which part of interconversion takes place in the free (unbound) form of the analyte and which part in the complexed (bound) form. Further, we propose a complete dynamic model of capillary electrophoresis of interconverting enantiomers based on solving a complete set of continuity equations for all constituents of the separation system together with complexation and acid-base equilibria. This allows a simulation of both linear and nonlinear mode of separation and understanding all processes taking place in such enantioseparation systems. We demonstrate the applicability of the method on determination of the rate constants of interconversion of oxazepam enantiomers separated in systems with charged cyclodextrin chiral selectors.     

Molecular interconversion behaviour in comprehensive two-dimensional gas chromatography

Comprehensive two-dimensional gas chromatography (GC x GC) is shown to provide information on dynamic molecular behaviour (interconversion), with the interconversion process occurring on both columns in the coupled-column experiment. The experiment requires suitable adjustment of both experimental conditions and relative dimensions of each of the columns. In this case, a longer column than normally employed in GC x GC allows sufficient retention duration on the second column, which permits the typical plateau-shape recognised for the interconversion process to be observed. The extent of interconversion depends on prevailing temperature, retention time, and the phase type. Polyethylene glycol-based phases were found to result in high interconversion kinetics, although terephthalic acid-terminated polyethylene glycol had a lesser extent of interconversion. Much less interconversion was seen for phenyl-methylpolysiloxane and cyclodextrin phases. This suggests that for the oximes, interconversion largely occurs in the stationary phase. Examples of different extents of interconversion in both dimensions are shown, including peak coalescence on the first column with little interconversion on the second column.     

Dynamic NMR studies of a potential chiroptical switch based on dithiocarbamate-iminodithiolane interconversion

[reaction: see text]. Variable temperature NMR spectra of the chiral spiro[(4-N,N-dimethyldithiocarbamato)-(2-N,N-dimethylimino)-1,3-dithiolane-5,9'-xanthene] show complex dynamics including degenerate interconversion of the dithiocarbamate and iminodithiolane groups. The rate of this switching process can be controlled by chemical modification: the analogous spiro[dithiolane-fluorene] derivative shows no interconversion. These novel materials have potential application as molecular switching elements in information storage devices.     

Ligand-induced interconversion of a coordination-organized porphyrin dimer: a potential fluorescence-based molecular memory monitor

A novel interconversion system between less-fluorescent stacked (S) dimer and fluorescent extended (E) dimer of the monoimidazolylbisporphyrinatozinc complex was investigated. The addition of pyridine induces transformation from the S to the E dimer, whereas the addition of acetic acid and subsequent heating reverses the transformation. The interconversion rate is controlled by ligand concentration and thermal treatment. The system can be applied to repeatedly readable molecular memory by highly sensitive fluorescence detection.     

Chemical relaxation studies on the system liver alcohol dehydrogenase, NADH and imidazole.

Several years ago, Theorell and Czerlinski conducted experiments on the system of horse liver alcohol dehydrogenase, reduced nicotinamide adenine dinucleotide and imidazole, using the first version of the temperature jump apparatus with detection of changes in fluorescence. These early experiments were repeated with improved instrumentation and confirmed the early experiments in general terms. However, the improved detection system allowed to measure a slight concentration dependence of the relaxation time of around 3 ms. Furthermore, the chemical relaxation time was smaller than the one determined earlier (by factor 2). The data were evaluated much more rigorously than before, allowing an appropriate interpretation of the results. The observed relaxation time is largely due to rate constants in an interconversion of ternary complexes, which are faster than three (of the four) dissociation rate constants, determined previously by Theorell and McKinley-McKee.1,2 This fact contributed to earlier difficulties of finding any concentration dependence. However, the binding of imidazole to the binary enzyme-coenzyme complex can be made to couple kinetically into the interconversion rate of the two ternary complexes. The observed signal derives largely from the ternary complex(es). A substantial fluorescence signal change is associated with the observed relaxation process, suggesting a relocation of the imidazole in reference to the nicotinamide moiety of the bound coenzyme. Nine models are considered with two types of coupling of pre-equilibria (none-all). Quantitative evaluations favor the model with two ternary complexes connected by an interconversion outside the four-step (bimolecular) cycle. The ternary complex outside the cycle has much higher fluorescence yield than the one inside. The interconversion equilibrium is near unity for imidazole. If it would be shifted very much to the side of the "dead-end" complex (as in isobutyramide?!), stimulating action could not take place.     

Gas chromatographic determination of the interconversion energy barrier for dialkyl 2,3-pentadienedioate enantiomers

The enantiomers of dialkyl 2,3-pentadienedioate undergo interconversion during gas chromatographic separation on chiral stationary phases. In this paper the on-column apparent interconversion kinetic and thermodynamic activation data were determined for dimethyl, diethyl, propylbutyl and dibutyl 2,3-pentadienedioate enantiomers by gas chromatographic separation of the racemic mixtures on a capillary column containing a polydimethylsiloxane stationary phase coupled to 2,3-di-O-methyl-6-O-tertbutyldimethylsilyl-beta-cyclodextrin. A deconvolution method was used to determine the individual enantiomer peak areas and retention times that are needed to calculate the interconversion rate constants and the energy barriers. The apparent rate constants and interconversion energy barriers decrease slightly with an increase in the alkyl chain length of the dialkyl 2,3-pentadienedioate esters. The optimum conformation of the dialkyl 2,3-pentadienedioate molecules, their separation selectivity factors and apparent interconversion enthalpy and entropy data changes with the alkyl chain length. The dependence of the apparent interconversion energy barrier (deltaG(app)(a-->b), deltaG(app)(b-->a)) on temperature was used to determine the apparent activation enthalpy (deltaH(app)(a-->b), deltaH(app)(b-->a)) and apparent entropy (deltaS(app)(a-->b), deltaS(app)(a-->b)) (where a denotes the first and b second eluted enantiomer). The comparison of the activation enthalpy and entropy (deltaS(app)(a-->b), deltaS(app)(a-->b)) indicated that the interconversion of dialkyl 2,3-pentadienedioate enantiomers on the HP-5+Chiraldex B-DM column series is an entropy driven process at 160 degrees C. Data obtained for dimethyl 2,3-pentadienedioate enantiomers on the HP-5+Chiraldex B-DM column series at 120 degrees C (deltaG(app)(a-->b) = 123.3 and deltaG(app)(b-->a) = 124.4 kJ mol(-1)) corresponds (at the 95% confidence interval) with the value of deltaG(#) = 128+/-1 kJ mol(-1) found at this temperature by gas chromatography using a two-dimensional stop flow technique on an empty capillary column [V. Schurig, F. Keller, S. Reich, M. Fluck, Tetrahedron: Asymmetry 8 (1997) 3475].     

Temperature effects on packed-capillary liquid chromatography of the X-ray contrast agent iodixanol

The effect of varying the operating temperature from 6 to 90 degrees C on the chromatographic performance of the exo-exo and exo-endo isomers of the X-ray contrast agent lodixanol in packed-capillary reversed-phase liquid chromatography shows increasing interconversion rates between the two isomeric conformers with increasing temperature. At 90 degrees C, Iodixanol elutes as one sharp peak due to an increased interconversion rate between the two isomeric conformers. Consequently, increased sensitivity is achieved. Temperature programming from 6 to 40 degrees C is utilized to optimize the resolution and determination of the exo-exo and exo-endo isomers. Temperature programming provides a significant decrease in the retention times in comparison with the isothermal separations while still preserving baseline separation of the isomers.     

Aminolysis reaction of calix [ 4 ] arene esters and crystal structures and conformational behaviors of calix[4]arene amides

We first make use of aminolysis of calix[4]arene esters to synthesize calix[4]arene amides. When the two ethyl esters of the calix[4]arene esters are aminolysized, the 1, 3-amide derivative is formed selectively. The crystal structures of the calix-[4]arene with two butyl amide (3b) and four butyl amide moieties (4b) were determined. The intermolecular hydrogen bonds make 4b form two-dimensional net work insolid state. The 1H NMR spectra prove that 3b is of a pinched cone conformation, while 4b and tetraheptylamide-calix[4]arene (6b) take fast interconversion between two C2v isomers in solution and appear an apparent cone conformation at room temperature. As decreasing temperature, the interconversion rate decreases gradually and, finally, the interconversion process is frozen at Tc = -10℃, which makes both conformations of 4b and 6b the pinched cone structures. The hydrogen bond improves the interconversion barrier, and the large different values of the potential barrier between 6b and 4b (or 6b) may     

X-ray diffraction analysis of nonequilibrium states in crystals: observation of an unstable conformer in flash-cooled crystals

Some molecules with a molecular skeleton similar to that of stilbenes and azobenzenes show orientational disorder in the crystals. If the disorder is dynamic, a conformational interconversion takes place through a pedal motion. In this study X-ray diffraction analyses of (E)-stilbene (1) and azobenzene (2) were carried out at various temperatures. We succeeded in observing thermodynamic nonequilibrium states that were generated by fast freezing of the conformational interconversion at low temperatures. The populations of the two conformers in crystals of 1 at 90 K varied with the cooling rate. Flash cooling of a crystal of 2 from room temperature to 90 K made it possible to observe the minor unstable conformer that does not exist in the equilibrium state at the same temperature.     

Structure and energetics of cyclopropane carboxaldehyde

We use the energies obtained by a focal point analysis including extrapolation from results with basis sets cc‐pVnZ and aug‐cc‐pVnZ with n up to 4 and correlation corrections through CCSD(T), to estimate thermodynamic functions for the syn and anti isomers of cyclopropane carboxaldehyde (CPCA). These agree with values obtained by well‐established thermochemical schemes CBS‐QB3 and G4. The structures obtained in these studies also conform to the best experimental determination of the rotational constants in the gas phase. The kinetics of gas phase interconversion of the syn‐ and anti‐isomers of CPCA have been studied by a chirped‐pulse dynamic rotational spectroscopy. Computational modeling of the internal rotational potential allows the estimate of the interconversion rates by statistical (RRKM) methods. RRKM rates using a range of barrier heights including a CBS‐Q estimate are more than 10× the rates deduced from the dynamical rotational spectra. This suggests that nonstatistical effects may be limiting the rate. Detailed study of the interconversion potential by a variant of the focal point analysis suggests that previous estimates of the barrier may be too low, and thus, the inferred rice‐ramsperger‐kassel‐marcus (RRKM) rate could be too high. These results cast some doubt on the presence of nonstatistical effects and suggest that molecular dynamics studies should be conducted to characterize the energy flow in detail. © 2013 Wiley Periodicals, Inc.     

Separation of triphenyl atropisomers of a pharmaceutical compound on a novel mixed mode stationary phase: a case study involving dynamic chromatography, dynamic NMR and molecular modeling

Analysis of atropisomers is of considerable interest in the pharmaceutical industry. For complex chiral molecules with several chiral centers hindered axial rotation can lead to formation of interconverting diastereomers that should be separable on achiral stationary phases. However, achieving the actual separation may be difficult as the on-column separation speed must match or be faster then the rate of isomer interconversion. Often, this requirement can be satisfied by using low-temperature conditions and by improving selectivity via use of chiral stationary phases. In the current study, we present an alternative approach utilizing an Obelisc R column, a novel mixed mode stationary phase that provided acceptable separation of triphenyl atropisomers inside a conventional HPLC temperature range. The separation was investigated under various chromatographic conditions. The interconversion chromatograms exhibited classic peak-plateau-peak behavior indicating the simultaneous atropisomer separation and interconversion. The elution profiles were integrated in order to deconvolute the peak areas of the "pure" (non-exchanged) and interconverted species; these data were used to obtain kinetic information. Analysis of retention data rendered thermodynamic information on the mechanism of retention and selectivity. Chromatographic kinetic data were complemented with variable-temperature NMR and molecular modeling studies, which provided additional support and insights into the energetics of the interconversion process.     

Isomerism of [64Cu-NOTA-Bn]-labeled radiotracers: separation of two complex isomers and determination of their interconversion energy barrier using ion pair chromatography

The model complex [(64)Cu((S)-p-NH(2)-Bn-NOTA)](-) ([(64)Cu]1) was used to study the isomerism of [(64)Cu-NOTA-Bn]-labeled radiotracers. Two complex isomers [(64)Cu]1a and [(64)Cu]1b, which were formed at a ratio of 1:9 during the complexation of [(64)Cu]Cu(2+) with (S)-p-NH(2)-Bn-NOTA, were separated using ion pair chromatography. To study the interconversion, the nonradioactive complex isomers Cu1a and Cu1b were separated and thermally treated at 90 °C in both ammonium acetate solution and deionized water. A faster interconversion rate was observed for both isomers with lower concentrations of ammonium ions. At the end of reaction, the thermodynamic Cu1a to Cu1b equilibrium ratio was 6:94. The particular energy barriers of the interconversion for Cu1a and Cu1b were 130 kJ mol(-1) and 140 kJ mol(-1). Spectrophotometric measurements with Cu1a and Cu1b revealed two isomers adopting different geometrical configurations.     

Radiation-induced cis–trans isomerization of polyisoprenes and temperature dependence of the equilibria

The cis-trans interconversion of polyisoprenes in solutions induced by γ-radiation in the presence of a sensitizer, which is any one of organic bromides or n-butyl mercaptan, was studied by using hevea and gutta percha as starting substances. The percentage cis remaining or converted after irradiation were determined by the infrared absorption. The equilibrium constants for the interconversion at 22, 60, and 100°C. were found to be 3.00, 5.25, and 7.33, respectively. The first-order rate constants for cis → trans and trans → cis isomerizations at 22°C. were calculated to be 9.05 and 2.91, respectively. The results were interpreted by the mechanism proposed by Golub, according to which the double bonds from π complexes with radiolytic fragments from sensitizers give a radical transition state capable of interconversion. However, our results showing that heating shifts the equilibrium toward trans isomer are not in accord with the mechanisms of the radiation-induced isomerization of polybutadiene of Golub and those for photoisomerization of aromatic azo compounds.     

Shape and spin state in four-coordinate transition-metal complexes: the case of the d(6) configuration

Generalized polyhedral interconversion coordinates are defined within the framework of Avnir's continuous shape measures. The application of such interconversion coordinates to the study of the potential energy surfaces that define the stereochemical choice in four-coordinate transition metal complexes with different spin states is presented, and the correlation between potential energy curves and distribution of experimental structures along the tetrahedron to square interconversion path is shown for the case of the d(6) transition-metal complexes.     

Collision-induced conformational changes in glycine

We present quantum dynamical calculations on the conformational changes of glycine in collisions with the He, Ne, and Ar rare-gas atoms. For two conformer interconversion processes (III-->I and IV-->I), we find that the probability of interconversion is dependent on several factors, including the energy of the collision, the angle at which the colliding atom approaches the glycine molecule, and the strength of the glycine-atom interaction. Furthermore, we show that attractive interactions between the colliding atom and the glycine molecule catalyze conformer interconversion at low collision energies. In previous infrared spectroscopy studies of glycine trapped in rare-gas matrices and helium clusters, conformer III has been consistently observed, but conformer IV has yet to be conclusively detected. Because of the calculated thermodynamic stability of conformer IV, its elusiveness has been attributed to the IV-->I conformer interconversion process. However, our calculations present little indication that IV-->I interconversion occurs more readily than III-->I interconversion. Although we cannot determine whether conformer IV interconverts during experimental Ne- and Ar-matrix depositions, our evidence suggests that the conformer should be present in helium droplets. Anharmonic vibrational frequency calculations illustrate that previous efforts to detect conformer IV may have been hindered by the overlap of its IR-absorption bands with those of other conformers. We propose that the redshifted symmetric -CH(2) stretch of conformer IV provides a means for its conclusive experimental detection.