Potential and performance of anisotropic 19F NMR for the enantiomeric analysis of fluorinated chiral active pharmaceutical ingredients.

Controlling the enantiomeric purity of chiral drugs is of paramount importance in pharmaceutical chemistry. Isotropic 1H NMR spectroscopy involving chiral agents is a widely used method for discriminating enantiomers and quantifying their relative proportions. However, the relatively weak spectral separation of enantiomers (1H Δδiso(R, S)) in frequency units at low and moderate magnetic fields, as well as the lack of versatility of a majority of those agents with respect to different chemical functions, may limit the general use of this approach. In this article, we investigate the analytical potential of 19F NMR in anisotropic chiral media for the enantiomeric analysis of fluorinated active pharmaceutical ingredients (API) via two residual anisotropic NMR interactions: the chemical shift anisotropy (19F-RCSA) and dipolar coupling ((19F-19F)-RDC). Lyotropic chiral liquid crystals (CLC) based on poly-γ-benzyl-L-glutamate (PBLG) show an interesting versatility and adaptability to enantiodiscrimination as illustrated for two chiral drugs, Flurbiprofen® (FLU) and Efavirenz® (EFA), which have very different chemical functions. The approach has been tested on a routine 300 MHz NMR spectrometer equipped with a standard probe (5 mm BBFO probe) in a high-throughput context (i.e., ≈10 s of NMR experiments) while the performance for enantiomeric excess (ee) measurement is evaluated in terms of trueness and precision. The limits of detection (LOD) determined were 0.17 and 0.16 µmol ml-1 for FLU and EFA, respectively, allow working in dilute conditions even with such a short experimental duration. The enantiodiscrimination capabilities are also discussed with respect to experimental features such as CLC composition and temperature.

Spatially Resolved Anisotropic Natural Abundance Deuterium 2D-NMR Spectroscopy Using Bimesophasic Lyotropic Chiral Systems.

In this paper, we describe, for the first time, the combined and original use of spatially resolved anisotropic natural abundance deuterium (ANAD) 2D-NMR experiments and bimesophasic lyotropic chiral systems to extract two independent sets of anisotropic parameters such as 2H-RQCs from a single NMR sample. As a pioneering example, we focus on a mixture of immiscible polypeptides (PBLG) and polyacetylene helical polymers (L-MSP) dissolved in weakly polar organic solvents (chloroform). Nondeuterated (D)-(+)-camphor is used as a model chiral solute. By providing two series of 2H-RQCs, this new analytical approach paves the way for applications in 3D structure elucidation with increased reliability and also opens up original investigations in terms of spectral enantiomeric discriminations and mixing of helical polymers.

Anisotropic 1 H STD-NMR Spectroscopy: Exploration of Enantiomer-Polypeptide Interactions in Chiral Oriented Environments.

The front cover artwork is provided by Dr. Philippe Lesot's group (NMR in Oriented Media, ICMMO, UMR CNRS 8182) at Université Paris-Saclay, France. The image shows four pieces of a puzzle: the magnet of an NMR spectrometer, the principle of the 1 H STD-NMR experiment and the 3D helical structure of the poly-γ-benzyl-L-glutamate polymer leading to a chiral liquid-crystalline phase that discriminates the enantiomers of a model chiral solute (1-phenethyl alcohol). Putting these pieces of the puzzle together allows us to identify the hydrogen sites of each enantiomer interacting with the polypeptide side chain. These new outcomes are a further step towards a global understanding of the chiral recognition that occurs in such media. Read the full text of the Research Article at 10.1002/cphc.202200508.

Anisotropic 1 H STD-NMR Spectroscopy: Exploration of Enantiomer-Polypeptide Interactions in Chiral Oriented Environments.

We explore and report for the first time the use of 1 H saturation transfer difference NMR experiments (STD-NMR) in weakly aligning chiral anisotropic media to identify the hydrogen sites of enantiomers of small chiral molecules interacting with the side-chain of poly-γ-benzyl-l-glutamate (PBLG), a helically chiral polypeptide polymer. The first experimental results obtained on three model mono-stereogenic compounds outcomes are highly promising and demonstrate the possibility to track down possible differences of spatial position of enantiomers at the vicinity of the polymer side-chain. Anisotropic STD experiments appear to be well suited for rapid screening of chiral analytes that bind favorably to orienting polymeric systems, while providing new insights into the mechanism of enantio-discrimination without resorting to the time-consuming determination of molecular order parameters.

Study and quantification of the enantiodiscrimination power of four polymeric chiral LLCs using NAD 2D-NMR.

Identifying and understanding the role of key molecular factors involved in the orientation/discrimination phenomena of analytes in polymer-based chiral liquid crystals (CLCs) are essential tasks for optimizing computational predictions (molecular dynamics simulation) of the existing orienting systems, as well as designing novel helically chiral polymers as new enantiodiscriminating aligning media. From this perspective, we propose to quantify and compare the enantiodiscrimination power of four homochiral polymer-based lyotropic liquid crystals (LLCs) toward a given chiral solute using their 2H residual quadrupolar couplings (2H-RQCs) measured by anisotropic natural abundance deuterium 2D-NMR (ANAD 2D-NMR). Two families of chiral polymers are investigated in this study: (i) poly-peptide polymers (PBLG and PCBLL), and (ii) polyacetylene polymers (PDA and L-MSP, a new system never published so far). As model solute, we investigate the case of camphor, an interesting rigid bicyclic chiral molecule possessing ten 2H-RQCs (10 inequivalent monodeurated isotopomers per enantiomer). In order to analyse the orientational behaviour of each enantiomer in a single oriented sample, while simplifying the identification of the (D/L)-isomer signals on spectra, a D-isomer enriched scalemic mixture (ee(D) = 30%) was used. Orientational data of camphor in each mesophase were calculated for the first time using the computer program ConArch+, modified to accept 2H-RQCs as anisotropic data input. Differences in enantiodiscriminations provided by the four aligning systems are examined and discussed in terms of structural and chemical features between polymers. The new L-MSP mesophase described in this work exhibits very promising enantiodiscrimination capacities.

Molecular enantiodiscrimination by NMR spectroscopy in chiral oriented systems: Concept, tools, and applications.

The study of enantiodiscriminations in relation to various facets of enantiomorphism (chirality/prochirality) and/or molecular symmetry is an exciting area of modern organic chemistry and an ongoing challenge for nuclear magnetic resonance (NMR) spectroscopists who have developed many useful analytical approaches to solve stereochemical problems. Among them, the anisotropic NMR using chiral aligning solvents has provided a set of new and original tools by making accessible all intramolecular, order-dependent NMR interactions (anisotropic interactions), such as residual chemical shift anisotropy (RCSA), residual dipolar coupling (RDC), and residual quadrupolar coupling (RQC) for spin I > 1/2, while preserving high spectral resolution. The force of NMR in enantiopure, oriented solvents lies on its ability to orient differently in average on the NMR timescale enantiomers of chiral molecules and enantiotopic elements of prochiral ones, leading distinct NMR spectra or signals to be detected. In this compendium mainly written for all chemists playing with (pro)chirality, we overview various key aspects of NMR in weakly aligning chiral solvents as the lyotropic liquid crystals (LLCs), in particular those developed in France to study (pro)chiral compounds in relation with chemists needs: study of enantiopurity of mixture, stereochemistry, natural isotopic fractionation, as well as molecular conformation and configuration. Key representative examples covering the diversity of enantiomorphism concept, and the main and most recent applications illustrating the analytical potential of this NMR in polypeptide-based chiral liquid crystals (CLCs) are examined. The latest analytical strategy developed to determine in-solution conformational distribution of flexibles solutes using NMR in polypeptide-based aligned solvents is also proposed.

Exploring the enantiomeric 13C position-specific isotope fractionation: challenges and anisotropic NMR-based analytical strategy.

Trying to answer the intriguing and fundamental question related to chiral induction/amplification at the origin of homochirality in Nature: "Is there a relationship between enantiomeric and isotopic fractionation of carbon 13 in chiral molecules?" is a difficult but stimulating challenge. Although isotropic 13C-PSIA NMR is a promising tool for the determination of (13C/12C) ratios capable of providing key 13C isotopic data for understanding the reaction mechanisms of biological processes or artificial transformations, this method does not provide access to any enantiomeric 13C isotopic data unless mirror-image isomers are first physically separated. Interestingly, 13C spectral enantiodiscriminations can be potentially performed in situ in the presence of enantiopure entities as chiral-europium complexes or chiral liquid crystals (CLCs). In this work, we explored for the first time the capabilities of the anisotropic 13C-{1H} NMR using PBLG-based lyotropic CLCs as enantiodiscriminating media in the context of the enantiomeric position-specific 13C isotope fractionation (EPSIF), within the requested precision of the order of the permil. As enantiomeric NMR signals are discriminated on the basis of a difference of 13C residual chemical shift anisotropy (13C-RCSA) prior to being deconvoluted, analysis of enantiomeric mixtures becomes possible. The analytical potential of this approach when using poly-γ-benzyl-L-glutamate (PBLG) is presented, and the preliminary quantitative results on small model chiral molecules obtained at 17.5 T with a cryogenic NMR probe are reported and discussed. A promising analytical approach based on anisotropic irm-13C-NMR spectrometry to potentially reveal the natural 13C/12C isotopic enantiofractionation effects in organic chiral molecules is proposed and discussed.

Chemical Synthesis of [2H]-Ethyl Tosylate and Exploration of Its Crypto-optically Active Character Combining Complementary Spectroscopic Tools.

Small chiral molecules are excellent candidates to push the boundaries of enantiodiscrimination analytical techniques. Here is reported the synthesis of two new deuterated chiral probes, (R)- and (S)-[2H]-ethyl tosylate, obtained with high enantiomeric excesses. Due to their crypto-optically active properties, the discrimination of each enantiomer is challenging. Whereas their enantiopurity is determined by 2H NMR in chiral anisotropic media, their identification was performed by combining quantum chemical calculations and vibrational circular dichroism analysis.

Deuterium Residual Quadrupolar Couplings: Crossing the Current Frontiers in the Relative Configuration Analysis of Natural Products.

The determination of the 3D structure (configuration and preferred conformation) of complex natural and synthetic organic molecules is a long-standing but still challenging task for chemists, with various implications in pharmaceutical sciences whether or not these substances have specific bioactivities. Nuclear magnetic resonance (NMR) in aligning media, either lyotropic liquid crystals (LLCs) or polymer gels, in combination with molecular modeling is a unique framework for solving complex structural problems whose analytical wealth lies in the establishment of nonlocal structural correlations. As an alternative to the already well-established anisotropic NMR parameters, such as RDCs (residual dipolar couplings) and RCSAs (residual chemical shift anisotropies), it is shown here that deuterium residual quadrupolar couplings (2H-RQCs) can be extracted from 2H 2D-NMR spectra recorded at the natural abundance level in samples oriented in a homopolypeptide LLCs (poly-γ-benzyl-l-glutamate (PBLG)). These 2H-RQCs were successfully used to address nontrivial structural problems in organic molecules. The performance and scope of this new tool is examined for two natural chiral compounds of pharmaceutical interest (strychnine and artemisinin). This is the first report in which the 3D structure/relative configuration of complex bioactive molecules is unambiguously determined using only 2H-RQCs, which, in this case, are at 2H natural abundance.

Tuning the Reactivity of a Heterogeneous Catalyst using N-Heterocyclic Carbene Ligands for C-H Activation Reactions.

We report the dramatic impact of the addition of N-heterocyclic carbenes (NHCs) on the reactivity and selectivity of heterogeneous Ru catalysts in the context of C-H activation reactions. Using a simple and robust method, we prepared a series of new air-stable catalysts starting from commercially available Ru on carbon (Ru/C) and differently substituted NHCs. Associated with C-H deuteration processes, depending on Ru/C-NHC ratios, the chemical outcome can be controlled to a large extent. Indeed, tuning the reactivity of the Ru catalyst with NHC enabled: 1) increased chemoselectivity and the regioselectivity for the deuteration of alcohols in organic media; 2) the synthesis of fragile pharmaceutically relevant deuterated heterocycles (azine, purine) that are otherwise completely reduced using unmodified commercial catalysts; 3) the discovery of a novel reactivity for such heterogeneous Ru catalysts, namely the selective C-1 deuteration of aldehydes.

2 H QUOSY 2D-NMR Experiments in Weakly Aligning Systems: From the Conventional to the Ultrafast Approach.

We describe three anisotropic ultrafast (UF) QUadrupolar Ordered SpectroscopY (QUOSY) 2D-NMR experiments (referred to as ADUF 2D NMR spectroscopy) designed for recording the 2 H homonuclear 2D spectra of weakly aligned (deuterated) solutes in sub-second experiment times. These new ADUF 2D experiments derive from the Q-COSY, Q-resolved and Q-DQ 2D pulse sequences (J. Am. Chem. Soc. 1999, 121, 5249) and allow the correlation between the two components of each quadrupolar doublet, and then their assignment on the basis of 2 H chemical shifts. The UF 2D pulse sequences are analyzed by using the Cartesian spin-operator formalism for spin I=1 nuclei with a small quadrupolar moment. The optimal experimental/practical conditions as well as the resolution, sensitivity and quantification issues of these ADUF 2D experiments are discussed on comparison to their conventional 2D counterparts and their analytical potentialities. Illustrative ADUF 2D experiments using deuterated achiral/prochiral/chiral solutes in poly-γ-benzyl-L-glutamate based chiral liquid crystals are presented, as well as the first examples of natural abundance deuterium (ANADUF) 2D spectrum using 14.1 T magnetic field and a basic gradient unit (53 G.cm-1 ) in oriented solvents.

Multinuclear NMR in polypeptide liquid crystals: Three fertile decades of methodological developments and analytical challenges.

NMR spectroscopy of oriented samples makes accessible residual anisotropic intramolecular NMR interactions, such as chemical shift anisotropy (RCSA), dipolar coupling (RDC), and quadrupolar coupling (RQC), while preserving high spectral resolution. In addition, in a chiral aligned environment, enantiomers of chiral molecules or enantiopic elements of prochiral compounds adopt different average orientations on the NMR timescale, and hence produce distinct NMR spectra or signals. NMR spectroscopy in chiral aligned media is a powerful analytical tool, and notably provides unique information on (pro)chirality analysis, natural isotopic fractionation, stereochemistry, as well as molecular conformation and configuration. Significant progress has been made in this area over the three last decades, particularly using polypeptide-based chiral liquid crystals (CLCs) made of organic solutions of helically chiral polymers (as PBLG) in organic solvents. This review presents an overview of NMR in polymeric LCs. In particular, we describe the theoretical tools and the major NMR methods that have been developed and applied to study (pro)chiral molecules dissolved in such oriented solvents. We also discuss the representative applications illustrating the analytical potential of this original NMR tool. This overview article is dedicated to thirty years of original contributions to the development of NMR spectroscopy in polypeptide-based chiral liquid crystals.

Hydrogen Isotope Exchange Catalyzed by Ru Nanocatalysts: Labelling of Complex Molecules Containing N-Heterocycles and Reaction Mechanism Insights.

Ruthenium nanocatalysis can provide effective deuteration and tritiation of oxazole, imidazole, triazole and carbazole substructures in complex molecules using D2 or T2 gas as isotopic sources. Depending on the substructure considered, this approach does not only represent a significant step forward in practice, with notably higher isotope uptakes, a broader substrate scope and a higher solvent applicability compared to existing procedures, but also the unique way to label important heterocycles using hydrogen isotope exchange. In terms of applications, the high incorporation of deuterium atoms, allows the synthesis of internal standards for LC-MS quantification. Moreover, the efficacy of the catalyst permits, even under subatmospheric pressure of T2 gas, the preparation of complex radiolabeled drugs owning high molar activities. From a fundamental point of view, a detailed DFT-based mechanistic study identifying undisclosed key intermediates, allowed a deeper understanding of C-H (and N-H) activation processes occurring at the surface of metallic nanoclusters.

2 H and 13 C NMR-Based Enantiodetection Using Polyacetylene versus Polypeptide Aligning Media: Versatile and Complementary Tools for Chemists.

In this work, the practical/analytical potential of an L-valine-derived polyacetylene (PLA) lyotropic liquid crystal (LLC) is examined to spectrally discriminate enantiomers (racemic mixture) or enantiotopic directions of a large collection (23) of (pro)chiral model compounds (from rigid to flexible and polar to apolar ones), thus covering various important aspects of enantiomorphism. Experimental 2 H-{1 H} (deuterated analytes and at natural abundance level) and 13 C-{1 H} NMR results are discussed in terms of the difference of 2 H-RQCs or 13 C-RCSAs and compared to those obtained in polypeptide-type LLCs (PBLG). The analysis of the NMR results provides an overview of the enantiodifferentiation capabilities of PLA and gives useful/practical hints for the chemist to select the most appropriate chiral oriented system. Astonishing NAD NMR results were obtained in the case of one of the simplest, chiral alkanes, 3-methylhexane. From a theoretical viewpoint, the data collected highlight the key molecular factors involved in orientation/discrimination processes, as a basis for optimizing computational prediction (molecular dynamics simulation), as well as designing novel helically chiral polymers as new enantiodiscriminating aligning media. In addition, a new, robust and efficient protocol to synthesize PLA and its enantiomer (PDA) on a large scale and with small polydispersities is proposed.

2 H and 13 C NMR-Based Enantiodetection Using Polyacetylene versus Polypeptide Aligning Media: Versatile and Complementary Tools for Chemists.

Invited for this month's cover are the collaborating groups of Dr. Philippe Lesot (DR CNRS) at Université Paris-Sud/Université Paris-Saclay, France, and Professor Michael Reggelin at TU Darmstadt, Germany. The cover shows the proton-decoupled natural abundance deuterium (NAD-{1 H}) Q-resolved Fz 2D-NMR spectrum of (±)-3-methylhexane measured in the anisotropic lyotropic chiral liquid-crystalline phase formed by a concentrated solution of a helically chiral polyarylacetylene in chloroform solvent. Seventeen out of the twenty possible deuterium quadrupolar doublets are observed, demonstrating the enormous enantiodifferentiating capability of the system, even for an unfunctionalized chiral alkane. Read the full text of the article at 10.1002/cplu.201800493.

Ruthenium-catalyzed hydrogen isotope exchange of C(sp3)-H bonds directed by a sulfur atom.

We present here the first example of C(sp3)-H activation directed by a sulfur atom. Based on this transformation catalyzed by Ru/C, we have developed a hydrogen isotope exchange reaction for the deuterium and tritium labelling of thioether substructures in complex molecules.

Integrated Computational Protocol for the Analysis of Quadrupolar Splittings from Natural-Abundance Deuterium NMR Spectra in (Chiral) Oriented Media.

Despite its low natural abundance, deuterium NMR spectroscopy in weakly oriented (chiral) solvents gives easy access to deuterium residual quadrupolar couplings (2 H-RQCs). These are formally equivalent to one-bond residual dipolar couplings ((13 C-1 H)-RDCs) for calculation of the Saupe tensor, and provide similar information for the study of molecular structure and orientational behavior. Because the quadrupolar interaction is one order of magnitude larger than the dipolar one, 2 H-RQC analysis is a much more sensitive tool for the detection of subtle structural differences and also tiny differences in molecular alignment, such as those observed for different enantiomers in chirally aligned media. To promote the analytical advantages of anisotropic, natural-abundance deuterium NMR (NAD NMR) spectroscopy in the organic chemistry community, we describe a 2 H-RQC/DFT-based integrated computational protocol for the evaluation of the order parameters of aligned solutes by using singular-value decomposition. Several examples of 2 H-RQC-assisted analysis of chiral and prochiral molecules dissolved in various polypeptide lyotropic chiral liquid crystals are reported. The role of the molecular shape in the ordering mechanism was investigated through the determination of intertensor angles between alignment tensors and inertia tensors by using the proposed protocol.

Investigation of Solute-Fiber Affinity and Orientational Ordering of Norbornadiene Interacting with Two-Polypeptide Chiral Liquid Crystalline Solvents by Natural Abundance Deuterium (NAD) NMR.

A prochiral bridged compound of C2v symmetry, the norbornadiene (NBD), oriented in a chiral liquid crystal composed of various mixtures of poly-γ-benzyl-l-glutamate (PBLG) and poly-ε-carboxy-l-lysine (PCBLL), two chiral homopolypeptides, is investigated using natural abundance deuterium 2D-NMR (NAD 2D-NMR) spectroscopy. In such chiral oriented solvents, enantiotopic directions are spectrally nonequivalent, and two distinct (2)H quadrupolar doublets associated with enantioisotopomeric pairs of NBD are detected. As the two homopolypeptides have the same absolute configuration but distinct chemical functions in their side chains, the variation of residual quadrupolar couplings (RQC's) allows the determination of the relative solute-fiber affinities toward the two polypeptides in these lyotropic bipolymeric systems. Besides the experimental measurement of RQC's and the determination of their signs at each inequivalent (2)H site, the elements of the second-rank order tensor, Sαß, are calculated by assuming a modeled structure. The variations of RQC's and diagonalized order parameters, Sα'α', are followed versus the relative proportion of two polypeptides in the chiral oriented mixture. The influence of the solute mass fraction in the two-homopolypeptide oriented samples is also examined as well as the case of homogeneous and uniform achiral mesophases "PBG-PCBL" made of two pairs of mirror-image homopolypeptides (PBLG/PBDG and PCBLL/PCBDL). In the latter, the solute ordering is modulated by the proportion of each type of homopolypeptide (chemical nature and absolute configuration), leading to eliminate the enantiodiscrimination mechanisms on the average. In the frame of a model, new insights on the solute-homopolypeptide fiber interactions are discussed.

Detection of quadrupolar nuclei by ultrafast 2D NMR: exploring the case of deuterated analytes aligned in chiral oriented solvents.

Anisotropic (2)H ultrafast (ADUF) 2D NMR spectroscopy for studying analytes dissolved in chiral liquid crystals (CLC) is investigated for the first time and the analytical possibilities of this method are evaluated. We demonstrate that these unconventional sub-second 2D experiments are compatible with the basic gradient units (40-60 G cm(-1)) that are implemented in routine spectrometers and allow the recording of (2)H signals of weakly aligned deuterated solutes in sub-second experimental times.

Synthesis and Stereochemical Assignment of Crypto-Optically Active (2) H6 -Neopentane.

The determination of the absolute configuration of chiral molecules is at the heart of asymmetric synthesis. Here we probe the spectroscopic limits for chiral discrimination with NMR spectroscopy in chiral aligned media and with vibrational circular dichroism spectroscopy of the sixfold-deuterated chiral neopentane. The study of this compound presents formidable challenges since its stereogenicity is only due to small mass differences. For this purpose, we selectively prepared both enantiomers of (2) H6 -1 through a concise synthesis utilizing multifunctional intermediates. While NMR spectroscopy in chiral aligned media could be used to characterize the precursors to (2) H6 -1, the final assignment could only be accomplished with VCD spectroscopy, despite the fleetingly small dichroic properties of 1. Both enantiomers were assigned by matching the VCD spectra with those computed with density functional theory.