Enantiomerization Rate Constants Research Articles

Investigation of the enantiomerization barriers of the phthalimidone derivatives EM12 and lenalidomide by dynamic electrokinetic chromatography.

The phthalimidone derivatives EM12 and lenalidomide, which are both structurally related to thalidomide, are highly interesting drugs and very recently lenalidomide attracted great attention as an antitumor and immune-modulating drug in the therapy for multiple myeloma. EM12 and lenalidomide are chiral, and the stereogenic carbon C-3 in the piperidine-2,6-dione moiety of these phthalimidone derivatives is prone to interconversion due to keto-enol tautomerization. The knowledge of the enantiomerization barrier is mandatory for pharmacokinetic studies and to develop a tailored therapy using the enantiopure or racemic drug. Here, we used dynamic EKC in combination with direct-calculation methods to determine the enantiomerization barriers of EM12 and lenalidomide. The separations of the enantiomers of EM12 and lenalidomide were performed in 50 mM aqueous disodium hydrogen phosphate buffer at pH 8 and 50 mM aqueous sodium tetraborate buffer at pH 9.3, respectively, using 20 mg/mL heptakis-(2,3-diacetyl-6-sulfato)-β-CD as a chiral additive. Enantiomerization of the compounds during the electrokinetic chromatographic separation resulted in pronounced plateau formation between the well-separated enantiomers. Peak form analysis of the experimentally obtained interconversion profiles yielded the enantiomerization rate constants k1 of EM12 and lenalidomide as well as the kinetic activation parameters ΔG(‡), ΔH(‡‡), and ΔS(‡) of enantiomerization by the evaluation of temperature-dependent measurements. The enantiomerization barrier ΔG(‡) was determined to be 98.3 ± 1.0 kJ/mol; the activation parameters ΔH(‡) = 46.1 ± 2.4 kJ/mol and ΔS(‡) = -170 ± 61 J/(K·mol) for EM12 and ΔG(‡) = 91.5 ± 1.0 kJ/mol, ΔH(‡) = 62.4 ± 5.4 kJ/mol, and ΔS(‡) = -98 ± 7 J/(K·mol) for lenalidomide. These findings were corroborated by density functional theory calculations at the B3LYP/3-21G level of theory of the ground state and intermediates considering an enantiomerization pathway via a keto-enol tautomerism.

Stereodynamics of Small 1,2‐Dialkyldiaziridines

Diaziridines are very interesting representatives of organic compounds containing stereogenic nitrogen atoms. In particular, 1,2-dialkyldiaziridines show extraordinarily high stereointegrity. The lone electron pairs of the nitrogen atoms are in trans configuration, avoiding a four-electron repulsive interaction. Furthermore, the trans configuration of the substituents at the nitrogen atoms is energetically favored because of reduced steric interactions. Therefore only two stereoisomers (enantiomers) are observed. At elevated temperatures the enantiomers are interconverting because of the limited stereointegrity of the chirotopic nitrogen atoms. The enantiomerization rate constants and the activation parameters of interconversion are of great interest. Here, we investigated the stereodynamics of a set of small 1,2-dialkyldiaziridines bearing short substituents (Me, Et, iPr, tBu), using enantioselective dynamic gas chromatography (DGC). Separation of enantiomers of all compounds, including the highly volatile 1,2-dimethyldiaziridine, was achieved using heptakis(2,3-di-O-ethyl-6-O-tert-butyldimethylsilyl)-β-cyclodextrin in 50% PS086 (w/w) as chiral stationary phase in fused silica capillaries with a length of up to 50 m. Measurements at variable temperatures were performed and reaction rate constants were determined using the unified equation of chromatography implemented in the software DCXplorer. The activation barriers at room temperature for 1-(tert-butyl)-2-ethyldiaziridine, ΔG(╪)(298K) = 123.8 kJ mol(-1) (ΔH(╪) = 115.5±2.9 kJ mol(-1), ΔS(╪) = -28±1 J mol(-1) K(-1)), and 1-ethyl-2-isopropyldiaziridine, ΔG(╪)(298K) = 124.2 kJ mol(-1) (ΔH(╪) = 113.1±2.4 kJ mol(-1), ΔS(╪) = -37±2 J mol(-1) K(-1)), were determined, representing some of the highest values observed for nitrogen inversion in diaziridines.

Kinetics of Aspartic Acid Isomerization and Enantiomerization in Model Aspartyl Tripeptides under Forced Conditions

The aim of the present study was the determination of the isomerization and enantiomerization of aspartic acid (Asp) in tripeptides. Capillary electrophoresis (CE) assays were developed and validated allowing the simultaneous determination of the diastereomeric alpha-D/L-Asp and beta-D/L-Asp peptides. Rapid isomerization and enantiomerization were noted for peptides with the Phe-Asp-GlyOH sequence at pH 10 and 80 degrees C while Gly-Asp-PheOH proved to be more stable due to the steric influence of the phenyl side chain. A kinetic model assuming a central role of the succinimide intermediate was used to fit the concentration versus time data. In incubations of L-Phe-alpha-L-Asp-GlyOH the ratio of alpha-Asp/beta-Asp peptides was about 1:4 in agreement with literature data. With regard to L-Asp and D-Asp peptides an alpha-Asp/beta-Asp ratio of about 1:3 and 1:5, respectively, was observed. The stereochemistry of Phe at the X-1 position affected the ratio of L-Asp/D-Asp implying an effect of the stereochemistry of neighboring amino acids on Asp enantiomerization. Modeling only overall Asp enantiomerization rate constants in accordance to literature data were observed for Asp peptides. In case of the asparagine (Asn) peptide the data could only be fitted to the models considering a direct conversion of L-Asn to a D-configured succinimide via an alternative pathway.

Simultaneous determination of enantiomerization and hydrolysis kinetic parameters of chiral N‐alkylbenzothiadiazine derivatives

On-column stopped flow multidimensional HPLC (sfMDHPLC) and dynamic high-performance liquid chromatography were applied to investigate the influence of alkyl substituents at the sulfonamidic and amino moieties of benzothiadiazine 1,1-dioxide derivatives on hydrolysis and enantiomerization rate constants. The data obtained indicate the presence of pyrrolo substituent at the 3,4 positions on benzothiadiazine rings inhibits the hydrolysis, whereas the enantiomerization occurs in acidic medium. Hydrolysis rates are quite similar for the two benzothiadiazines methyl substituted to nitrogen at 2- and 4-positions. Conversely, enantiomerization rate of 4-N-methyl substituted is significantly higher than 2-N-methyl substituted.

Determination of the interconversion energy barrier of 2,3‐pentadienedioic acid enantiomers by HPLC. 2. On‐column interconversion

In this paper, an HPLC method is used to determine the enantiomerization barrier of 2,3-pentadienedioic acid enantiomers. The racemate of 2,3-pentadienedioic acid was separated by HPLC on a chiral CHIROBIOTIC T column with a 90:10 (100:0.5:0.5 MeOH/HOAc/TEA)/H2O mobile phase. Peak areas of enantiomers prior to (A(+)0, A(-)0) and after the separation (A(+), A(-)), were used for calculation of the rate constants and the enantiomerization barrier, as determined by computer-assisted peak deconvolution of the peak clusters on the chromatograms. The kinetic equation for irreversible reactions was used to determine the apparent enantiomerization rate constants and the interconversion energy barrier. The dependence of the apparent enantiomerization barrier (deltaG1(app), deltaG-1(app)) on temperature was used to determine the apparent activation enthalpy (deltaH1(app), deltaH(-1)app) and entropy (deltaS1(app), deltaS-1(app)) for the interconversion of 2,3-pentadienedioic acid enantiomers, where the coefficients 1 and -1 designate the interconversions (+) --> (-) and (-) --> (+), respectively.

Fast and precise access to enantiomerization rate constants in dynamic chromatography

An analytical solution of the unified equation to evaluate elution profiles of interconverting enantiomers in dynamic chromatography is presented. Rate constants k1 and k(-1) and Gibbs activation energies are directly obtained from the chromatographic parameters (retention times tR A and tR A of the interconverting enantiomers, the peak widths at half height wA and wB, and the relative plateau height hp), and the initial amounts A0 and B0 of the enantiomers without any iterative and time consuming computational step. Therefore, this equation is no longer limited to racemic analytes. The analytical solution presented here was validated by comparison with a dataset of 125,000 simulated elution profiles of enantiomerizations. Furthermore, it was found that the recovery rate from a defined dataset is on average 40% higher using the unified equation compared to evaluation methods based on iterative computer simulation. The new equation was applied to determine the enantiomerization rate constant of 1-n-butyl-2-tert-butyldiaziridine by enantioselective gas chromatography. The activation parameters (DeltaH(double dagger) = 112.6 +/- 2.5 kJ/mol and DeltaS(double dagger) = -27 +/- 2 J/(K mol) were obtained from temperature-dependent measurements between 100 degrees C and 140 degrees C in 10K steps.

Static and Dynamic Stereochemistry of Tris(9-triptycyl)stannane Derivatives

Abstract Static and dynamic stereochemistry of several tris(9-triptycyl)stannane derivatives Tp3SnX (X = H, halogens, and alkyl groups) were studied. X-ray crystallography of the bromo and methyl compounds showed that the Sn atom was fundamentally tetrahedral and that the three Tp groups meshed with each other like bevel gears and formed a near-C3 chiral conformation. The rate constants for enantiomerization of the chiral conformation in solution could be obtained by the lineshape analysis of the aromatic proton signals, even when a direct probe for enantiomerization such as a benzyl group was absent. The energy barrier to enantiomerization decreased as the substituent X became bulkier, suggesting that the ground state is more destabilized than the transition state upon introduction of a bulkier group.

Separate Determination of Rate Constants from Reversible Reactions in a Chromatographic Column and Eluent Using Empirical Peak Shape Equations

A new mathematical‐analytical equation is derived for fitting purposes in the determination of mobile phase rate constants of enantiomerization from liquid chromatographic reactor experiments with a recycling of the internal chromatogram via an empty capillary. The equation was successfully tested with results from numerical calculations, as well as results from the on‐column enantiomerization of oxazepam.

Probing the stereointegrity of Tröger's base--a dynamic electrokinetic chromatographic study.

Under acidic conditions the enantiomers of Tröger's base 1 (2,8-dimethyl-6 H,12 H-5,11-methanodibenzo[b,f][1,5]diazozine) are subject to enantiomerization. During enantioselective dynamic electrokinetic chromatography using 10 mM hydroxypropyl-beta-cyclodextrin as the chiral mobile phase additive in 50 mM tris/phosphate buffer at pH 2.2, enantiomerization of Tröger's base gives rise to characteristic elution profiles featuring plateau formation and peak broadening. Introduction of a permanent positive charge attributed to quaternization in the monobenzylated derivative of Tröger's base 2 (5-benzyl-2,8-dimethyl-6 H,12 H-5,11-methanodibenzo[b,f][1,5]diazozinium bromide) decreases the enantiomerization barrier significantly. To determine the rate constants of enantiomerization the experimental chromatograms were evaluated by a direct calculation method and by using the computer simulation program ChromWin. From temperature-dependent measurements the Eyring activation parameters for 1 and 2 were determined: 1: DeltaG( not equal ) (298 K)=100.9+/-0.5 kJ mol(-1), DeltaH( not equal )=89.5+/-2.0 kJ mol(-1), DeltaS( not equal )=-42+/-10 J K(-1) mol(-1); 2: DeltaG( not equal ) (298 K)=90.2+/-0.5 kJ mol(-1), DeltaH( not equal )=91.4+/-2.0 kJ mol(-1), DeltaS( not equal )=9.8+/-10 J K(-1) mol(-1).

Determination of the enantiomerization energy barrier of some 3-hydroxy-1,4-benzodiazepine drugs by supercritical fluid chromatography

The first-order kinetic equation for irreversible reactions was used to determine the enantiomerization barrier of some of 3-hydroxy-1,4-benzodiazepine enantiomers by supercritical fluid chromatography (SFC). The racemates of lorazepam, oxazepam and temazepam were separated by SFC on chiral ( R, R)-Whelk-O1 column with supercritical carbon dioxide containing 12.5% methanol and 0.5% diethylamine as a mobile phase. Peak areas of enantiomers prior to ( A A0 , A B0 ) and after the separation ( A A , A B ), used for calculation of the enantiomerization barrier, were determined by computer-assisted peak deconvolution of peak clusters from the chromatograms. It was demonstrated for the first time that using a model for a four-peak cluster produces height precise results, and most closely approximates the published results. The kinetic equation for irreversible reactions was used to determine apparent enantiomerization rate constants. The dependence of the apparent enatiomerization barrier (Δ G app A→ B , Δ G app B→ A ) on temperature was used to determine apparent activation enthalpy (Δ H app R→ S , Δ H app S→ R ) and entropy (Δ S app R→ S , Δ S app S→ R ) for all studied benzodiazepines.

Approximation function for the direct calculation of rate constants and Gibbs activation energies of enantiomerization of racemic mixtures from chromatographic parameters in dynamic chromatography

An approximation function for enantioselective dynamic chromatography of racemic mixtures of interconverting enantiomers has been derived that allows the direct calculation of enantiomerization rate constants ( k 1 and k −1) and Gibbs activation energies of enantiomerization, Δ G #, from chromatographic parameters, i.e., retention times of the enantiomers A and B ( t R A and t R B), peak widths at half height ( w A and w B) and the relative plateau height ( h plateau), without computer simulation. The reaction rate constants of enantiomerization, k −1, obtained with this approximation function, have been validated by comparison with a simulated dataset of 15 625 chromatograms. The mean, standard deviation and confidence interval show a high correlation between the approximated and simulated rate constants. The average deviation from the Gibbs activation enthalpy of enantiomerization, Δ G #, has been estimated to be as small as about ±0.11 RT.

Parametric Studies on the Determination of Enantiomerization Rate Constants from Liquid Chromatographic Data by Empirical Peak Shape Equations for Multi-Step Consecutive Reactions

On-column enantiomerization is a well documented and practically important special case of the occurrence of a reversible reaction in a chromatographic reactor. The present paper explains the possibility to obtain enantiomerization rate constants from liquid chromatographic data using analytical equations derived for multi-step consecutive reactions. The results are compared with results from a finite difference method, and the application to experimental data is demonstrated.

Chiral 2‐Aryl‐2‐methyl‐2H‐1‐benzopyrans: Synthesis, characterization of enantiomers, and barriers to thermal racemization

AbstractThe ease of thermal breaking of the C(sp3)O bond of the 2‐aryl‐2‐methyl‐2H‐1‐benzopyrans 1–9 was evaluated by measuring the free energy (ΔG) of the racemization reaction of optically active compounds. The variation of ΔG of the thermal ring opening in terms of structural modifications is discussed. The synthesis of the studied compounds, the preparative separation of enantiomers by liquid chromatography, the determination of enantiomeric purity, the circular dichroism of enriched enantiomers, and the measurement of rate constants of enantiomerization by monitoring the decrease of the polarimetric angle of rotation at suitable temperatures are described.

Enantiomerization of an atropisomeric drug

The antipsoriatic 10-(3-chlorophenyl)-6,8,9,10-tetrahydrobenzo[b][1,8] naphthyridin-5(7H)-one, Sch 40120, is chiral only because it lacks planarity and possesses a stereogenic axis. It comprises short-lived, interconverting atropisomeric enantiomers distinguished by the chlorine substitutent. The atropisomers form diastereomeric complexes with the shift reagent (R)-(−)-2,2,2-trifluoro-1-(9-anthryl)ethanol, which were detected by 1H NMR spectroscopy. Liquid chromatography on an ovomucoid chiral column isolated each enantiomer from the racemic mixture. Re-injections of the separated enantiomers onto the same column held constant at 10°C established that each enantiomer formed the other. Under identical chromatographic conditions, both stereoisomers independently recreated the racemic mixture. The calculated enantiomer half-life lasted 1.6 min at the physiological temperature of 37°C. Simulations of dynamic liquid chromatograms acquired with a chiral stationary phase indirectly yielded values of the half-lives. The chromatograms were modeled with the computer program SIMUL. Also determined were the rate constants for enantiomerization and the corresponding Gibbs free energies of activation, all at varying temperatures. At 37°C, the rate constant and activation energy respectively equaled 0.213 min−1 and 21.6 kcal mole−1. An Arrhenius plot was linear. The intractably brief life spans necessitated development of the racemic drug, rather than advancement of one enantiomer only. The pharmacological, biological, and chemical consequences of molecular asymmetry inherent to the drug were therefore nil. © 1996 Wiley-Liss, Inc.

Investigation of the Enantiomerization Barrier of Homofuran by Computer Simulation of Interconversion Profiles Obtained by Complexation Gas Chromatography

AbstractBy computer simulation of experimental interconversion profiles, obtained by complexation gas chromatography, rate constants of enantiomerization have been determined for homofuran between 95 and 130°C. Since enantiomerization proceeds at similar rates in the mobile and stationary phase, the rate constants obtained by an Arrhenius plot are in excellent agreement with values determined independently by polarimetry between 60 and 90°C. We thus demonstrate that dynamic chromatography can be an easy means for the rapid determination of enantiomerization (or isomerization) barriers of ca. 70–120 kJ mol‐1 requiring only minute amounts of racemic sample.

Enantiomerization during HPLC on an optically active sorbent. Deconvolution of experimental chromatograms

The enantiomers of 1,3,7-trimethylbenzo[c]phenanthrene, a chiral helicene hydrocarbon, are separated on triacetyl-cellulose but interconvert during HPLC. Column temperatures were between 2 and 22°C and linear velocities between 0.3 and 1.2 mm s−1. Simultaneous photometric and polarimetric detection resulted in deconvoluted experimental chromatograms, showing the relative concentration of each of the enantiomers as aseparate function of the retention volume. The use of deconvoluted chromatograms for determining rate constants of enantiomerization is discussed. It is of interest that the rate constant at 22.3°C in solution, i.e. in the absence of a sorbent, has been obtained by an independent method: stopped-flow monitoring of racemization after HPLC on triacetylcellulose.