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NMR spectroscopy was used to investigate the binding of chiral compounds to chiral molecular micelles.  The surfactant monomer units in the molecular micelles contained either a chiral amino acid or a dipeptide headgroup.  The monomers were polymerized at the end of each surfactant's hydrocarbon chain.   These molecular micelles are used as chiral selectors in electrokinetic chromatography (EKC).  EKC is a modified form of capillary electrophoresis where the run buffer contains a modifier like a surfactant micelle, cyclodextrin, or polymer.  When the modifier is chiral, the enantiomers of chiral compounds may interact differentially with the modifier as they both move through the capillary.  These differential, stereoselective interactions between the enantiomers and modifier cause the enantiomers to move with different velocities, thus bringing about chiral separation.

NMR spectroscopy was used to investigate the intermolecular interactions leading to chiral recognition in three molecular micelle: chiral molecule mixtures.  In the first project, NMR was used to study the interactions of seven b-blocker drugs to molecular micelles containing either nine, ten, or eleven carbons in their hydrocarbon chain and a chiral carbonyl-L-leucinate headgroup.  NMR NOESY experiments were used to investigate the conformation of the surfactant headgroups.   NOE interactions between the leucine amino acid and hydrocarbon chain protons suggested that the surfactant headgroups formed a folded conformation creating a chiral pocket into which chiral molecules inserted.  This conclusion was confirmed by NOESY experiments with mixtures containing the same molecular micelles and the chiral binaphthyl compound 1,1'-binaphthyl-2,2'-diyl hydrogen phosphate (BNP).  These experiments showed that BNP protons near the molecule's phosphate group pointed toward the surfactant headgroup and protons on the opposite side of the molecule pointed toward the hydrocarbon chain.  This result confirmed that the surfactant formed a chiral pocket into which the b-blockers and other chiral compounds inserted.

NMR diffusion experiments were also used to investigate the thermodynamics underlying the interactions between the b-blockers and the molecular micelles.  Seven b-blockers with increasing hydrophobicities were included in this study.  The most hydrophilic b-blocker was atenolol and the most hydrophobic was propranolol.  The NMR diffusion experiments allowed the molecular micelle association constants and free energies of binding for each b-blocker to be measured.  The free energies of binding from NMR were found to scale linearly with EKC retention times from the literature.¹  This result confirmed that the b-blockers separate based upon differences in their free energies of binding to the molecular micelle.  The strongest and weakest linear correlations were found, respectively, for molecular micelles containing nine and eleven carbons in their hydrocarbon chains.  Furthermore, differences in the free energies of binding of (R) and (S)-atenolol enantiomers to the molecular micelles also correlated with EKC separation results.  The differences in the (R) and (S)-atenolol DG values for polymers with hydrocarbon chains containing ten and eleven carbons were greater than the differences for polymers containing eight or nine carbons.  In EKC separations, chiral resolution was highest in separations with molecular micelles containing ten or eleven carbon atoms.  This result confirms that chiral resolution in EKC results from differences in the free energies of binding of the chiral compound's enantiomers to the molecular micelle.

NMR spectroscopy was also used to investigate the interaction between enantiomers of the chiral the drugs ephedrine, pseudoephedrine, and norephedrine and micelles formed from the chiral surfactant N-dodecocycarbonylvaline (DDCV).  The chiral surfactant is sold by Waters, Inc. as Enantioselect.    NMR diffusion experiments were used to establish the association constants and free energies of binding of each ephedrine enantiomer to (S) and (R) enantiomers of the surfactant micelles.  These experiments showed that ephedrine exhibited stronger binding to the micelles than the pseudoephedrine or norephedrine, while the norephedrine: DDCV association constants spanned a wider range than ephedrine or pseudoephedrine.    Two-dimensional ROESY experiments were conducted to probe the intermolecular interactions between the ephedrines and micelles. The ROESY data suggested that one strong H-bond between the ephedrine molecules and the micelle produced a complex with a more negative free energy of binding than complexes where two weaker and/or two competing H-bonds were formed.  The ROESY spectra also showed that when the ephedrines bound to the micelles, the drugs remained near the micelle surface within a chiral pocket, pointing their aromatic rings towards the hydrocarbon chain.  Little interaction of the ephedrines with the hydrophobic core of the micelle was observed.

            Finally, the NMR relaxation rate ratio method was used to investigate the binding of the chiral compound BNP to six molecular micelles with different dipeptide headgroups.  Spin-lattice and spin-spin relaxation times were measured for each of the BNP protons and those values were in turn used to calculate each BNP proton's correlation time or the time required for the proton to move through one radian.²  These experiments showed that in the presence of molecular micelles such as poly-(sodium undecyl-(L,L)-valine-leucine) (poly-(L,L)-SUVL)  that gave poor chiral resolution, all BNP protons had the same correlation time.  Thus the BNP molecule moved in an isotropic fashion and each part of the molecule experienced similar interactions with the polymer.  On the other hand, in experiments with micelles such as poly-(sodium undecyl-(L,L)-leucine-valine) (poly-(L,L)-SULV)which gave high chiral resolution, the BNP protons had different correlation times.  For example, in experiments with poly(SULV) correlation times ranged from 1.92±0.06 to 3.38±0.06  ns.  This result showed that in the presences of poly(SULV), the BNP molecule moved in an anisotropic fashion caused by different portions of the molecule experiencing different interactions with the polymer.  Thus, the motional anisotropy of bound chiral molecules was found to be an effective probe of chiral resolution. 

            These experiments taken together show the utility of NMR spectroscopy in investigating chiral recognition.  NOESY and ROESY experiments report on the structures of the intermolecular complexes, NMR diffusion studies investigate the thermodynamics of the chiral compound: molecular micelle association, and NMR relaxation experiments report on the motional dynamics of the bound chiral molecules.

1.     Ali Rizvi, S.A.; Shamsi, S.A. Electrophoresis 2005, 26, 4172–4186.

2.     Carper, W.R.; Keller, C.E. J. Phys. Chem. A. 1997, 101, 3246-3250.