NMR enantiodifferentiation studies are greatly improved by the simultaneous determination of ¹H and ¹³C chemical shift differences through the analysis of highly resolved cross-peaks in spectral aliased pure shift (SAPS) HSQC spectra. The determination of enantiomeric purity can be accomplished by NMR spectroscopy using a great variety of auxiliary chiral sources. Of these, chiral solvating agents (CSAs), such as the so-called Pirkle alcohol (PA) or cyclodextrins (CDs), have been widely used. They do not typically introduce significant line-broadening, the sample is easily prepared and the analysis is quickly performed by observing chemical shift differences (ΔΔδ) between the resulting diasteromeric complexes in conventional ¹H NMR spectra. However, signal enantiodifferentiation using CSAs is not uniform for all protons and in many cases, low ΔΔδ values and signal overlap caused by complex multiplets lead to the lack of spectral signal dispersion that preclude a straightforward analysis. Alternatively, enantiodifferentiation using ¹³C NMR spectroscopy can be more advantageous because singlet signals are analyzed, although its routine use is limited by its low sensitivity.

A frequency-selective 1D ¹H nuclear magnetic resonance (NMR) experiment for the fast and sensitive determination of chemical-shift differences between overlapped resonances is proposed. The resulting fully homodecoupled ¹H NMR resonances appear as resolved 1D singlets without their typical J(HH) coupling constant multiplet structures. The high signal dispersion that is achieved is then exploited in enantiodiscrimination studies by using chiral solvating agents.

The utility of ¹³C NMR spectroscopy for the differentiation of enantiomers using chiral solvating agents (CSA) is stated. The intrinsic high dispersion of ¹³C nucleus, as well as the singlet nature of the signals in standard experiments makes ¹³C NMR experiments a powerful alternative or complement to ¹H NMR experiments; specially, when studying pure compounds with complex proton spectra or mixtures of compounds, as in chiral metaboLomics, where severe overlapping exists in proton spectrum. To evaluate and compare the quality of the enantioresolution of a NMR signal we introduce the enantiodifferentiation quotient, E, that considers the complexity of ¹H multiplets (and in general the width) of the original signal. It has been observed that the error in the measurement of the enantiomeric molar ratio can be related to the E value.

Differences in molecular chirality remain an important issue in drug metabolism and pharmacokinetics for the pharmaceutical industry and regulatory authorities, and chirality is an important feature of many endogenous metabolites. We present a method for the rapid, direct differentiation and identification of chiral drug enantiomers in human urine without pretreatment of any kind. Using the well-known anti-inflammatory chemical ibuprofen as one example, we demonstrate that the enantiomers of ibuprofen and the diastereoisomers of one of its main metabolites, the glucuronidated carboxylate derivative, can be resolved by ¹H NMR spectroscopy as a consequence of direct addition of the chiral cosolvating agent (CSA) β-cyclodextrin (βCD). This approach is simple, rapid, and robust, involves minimal sample manipulation, and does not require derivatization or purification of the sample. In addition, the method should allow the enantiodifferentiation of endogenous chiral metabolites, and this has potential value for differentiating metabolites from mammalian and microbial sources in biofluids. From these initial findings, we propose that more extensive and detailed enantiospecific metabolic profiling could be possible using CSA-NMR spectroscopy than has been previously reported.

The determination of the enantiomeric purity of drugs and their intermediates has now become of crucial importance. Most of the methods applied in determining the enantiomeric excess (ee) of chiral compounds require labour-intensive steps, which involve derivatisation and purification of the product, with the possibility of a kinetic resolution as an important drawback. The majority of these methods are chromatographic and mass spectrometry-based techniques. NMR methods developed for this purpose have recently undergone remarkable advances. We have recently developed two chiral solvating agents (CSA) capable of enantiodifferentiating a wide variety of compounds, such as amines, alcohols, epoxides and products with and without aryl groups. The advantages of these auxiliaries are firstly, that no derivatisation or purification is required (the enantiodifferentiation occurs via noncovalent interactions) and secondly, if required, the sample can be easily recovered by chromatography. Furthermore, the drawbacks of kinetic resolution are avoided.

Several alcohols containing anthracene and trifluoromethyl groups, such as Pirkle's alcohol¹ or α,α^′-bis(trifluoromethyl)-9,10-anthracenedimethanol,² which are commercially available, are used as chiral solvating agents (CSA). The presence of a bulky group, such as tert-butyl ³ or adamantyl, transmits structural rigidity that compensates for the lack of acidity in the methinic proton. The presence of the aromatic group (anthracene) in the structure is one of the factors that allows the CSA to differentiate between enantiomers. The per-fluorophenyl group has a strong electron withdrawing character and its presence in the CSA could increase the zones around the enantiomers where the magnetic field of the NMR becomes modified by the presence of the anisotropic groups. The preparation of the enantiomers of 9-anthrylpentafluorophenylmethanol 1 gives us a novel CSA with new interactions and new associations that offer new possibilities for enantiodistinction. The synthesis, structure and behaviour as chiral solvating agents of the enantiomers of 9-anthrylpentafluorophenylmethanol is reported.

The preparation of the enantiomers of α,α′-bis(trifluoromethyl)-1,8-anthracenedimethanol (ABTE-18) is described, and their conformational behaviour studied. These enantiomers are very active when used as chiral solvating agents in the presence of several compounds. This compound conserves the capacity to interact with substrates, forming two hydrogen bonds at the same time. In addition, the accessibility of carbon atom 10 indicates that this compound might have other future applications.