Absolute configurations of synthetic molecular scaffolds from

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Perspective

Absolute configurations of synthetic molecular scaffolds from vibrational CD spectroscopy Christian Merten, Tino P. Golub, and Nora M. Kreienborg J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.9b00466 • Publication Date (Web): 02 May 2019 Downloaded from http://pubs.acs.org on May 3, 2019

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The Journal of Organic Chemistry

Absolute configurations of synthetic molecular scaffolds from vibrational CD spectroscopy Christian Merten*, Tino P. Golub, Nora M. Kreienborg Ruhr Universität Bochum, Organische Chemie II, Universitätsstraße 150, 44780 Bochum, Germany. [email protected]. www.mertenlab.de

ABSTRACT

Vibrational circular dichroism (VCD) spectroscopy is one of the most powerful techniques for the determination of absolute configurations (AC), as it does not require any specific UV/Vis chromophores, no chemical derivatization and no growth of suitable crystals. In the last decade it has become increasingly recognized by chemists from various fields of synthetic chemistry such as total synthesis and drug discovery as well as from developers of asymmetric catalysts. This review gives an overview about the most important experimental aspects of a VCD-based AC determination and explains the theoretical analysis. The comparison of experimental and

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computational spectra that leads to the final conclusion about the AC of the target molecules is described. In addition, the review summarizes unique VCD studies carried out in the period 2008-2018 that focus on the determination of unknown ACs of new compounds, which were obtained in its enantiopure form either through direct asymmetric synthesis or chiral chromatography.

INTRODUCTION As the extension of the more widely known electronic circular dichroism (ECD) spectroscopy, vibrational circular dichroism (VCD) measures the differential absorbance of left- and rightcircularly polarized (LCP/RCP) light during vibrational transitions and can thus be considered the chiral version of IR spectroscopy. The VCD spectrum of a given enantiomer of a chiral molecule shows either a positive or a negative band for each of the vibrational modes, depending on the preferential absorbance of either type of circularly polarized light. Similarly, the opposite enantiomer will feature a mirror imaged spectrum with all bands having opposite signs. By comparison with computationally predicted IR and VCD spectra, the spectral signatures can be used to assign the AC of the sample with very high confidence. The maximum in molecular size and complexity which can still be characterized by VCD spectroscopy is thus mainly determined by computational feasibility of the corresponding spectra calculations. The currently investigated molecular size range already spans from rather small natural products1 and synthetic molecules like those covered in this review article to polymers2-3 and supramolecular assemblies.4-6 In addition, being based on IR spectroscopy, VCD spectra have been shown to be very sensitive to intermolecular interactions7 and have thus been used to study, for instance, reactant-catalyst binding and chirality transfer in catalysis.8-9


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There are several practical advantages of VCD spectroscopy, which make it a very universal method for AC determinations. Unlike in ECD spectroscopy, where special chromophores are required to generate or enhance the chiroptical response, all chiral molecules are IR active and hence will show a VCD spectrum. Therefore, there is no need to chemically modify the compound, which is necessary for the application of NMR-based methods such the Mosher’s ester procedure.10 Moreover, VCD spectroscopy enables access to the ACs of compounds which cannot be crystallized, i.e. liquids and oily substances, or for which suitable crystals cannot be grown. Nevertheless, while VCD spectroscopy has become an important tool in natural products chemistry,1 in particular x-ray crystallography and NMR spectroscopy are currently the most frequently used methods for the AC determination of new synthetic scaffolds in academia.

Scheme 1. Examples for AC assignments from pharmaceutical industry using VCD spectroscopy.11-13

In contrast, the pharmaceutical industry has accepted VCD spectroscopy as their preferred choice for AC assignments. For a high-throughput industry, crystallography-based assignments are often too slow because of the aforementioned additional modifications that are required or simply because of the time (and luck) required to obtain suitable single crystals for analysis. There is, however, not much publically accessible proof for its popularity,14 as most of the characterized compounds are synthesis intermediates and/or proprietary, and hence will never be



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published. A few examples, which have been reported, are shown in Scheme 1. Instead of publication output, the importance of VCD for the pharmaceutical industry can be measured in instrument sales numbers (nearly every major pharmaceutical company has access to a VCD instrument) or the number of patents, which cite VCD as main method for AC assignments (currently more than 130 applications and 95 patents granted).15-16 In addition, since December 2016, VCD spectroscopy is recognized as standard method for AC assignments in the US Pharmacopeia (USP 39-NF34, Chapters 728 and 1728). Overall, for the last decade covered by this review article, we estimate the number of AC assignments in pharmaceutical industry using VCD spectroscopy to be in the thousands. The aim of this perspective article is not only to summarize the AC determinations of synthetic chiral molecules published over the last decade. Moreover, we want to use this collection of examples to showcase the diversity of structures accessible with VCD spectroscopy and thereby encourage chemists to consider VCD spectroscopy as powerful, fast and reliable method for the characterization of stereochemistry. Using examples from our own work, we also explain the typical procedure of an AC assignment using VCD spectroscopy in order to make it transparent and more easily comprehensible for readers new to the field and to those who read or co-author any organic or inorganic journal article that makes use of VCD.

GENERAL PROCEDURE FOR AC DETERMINATION USING VCD SPECTROSCOPY

In the spirit of this review and without going into too much technical detail, this section briefly summarizes some important aspects regarding experimental requirements and computational approaches for a VCD spectral analysis. We also discuss the final comparison of experimental


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and predicted IR and VCD spectra resulting in the assignment of an AC. Intentionally, we leave out descriptions of the spectroscopic instrumentation or the theoretical foundation of VCD and refer the interested reader to monographs17-18 and other reviews dealing with these topics.15, 19 Instead, the purpose of this section is rather to outline practical aspects of a VCD spectral analysis and thus to provide a solid basis of information for those getting started with or those seeking help from VCD spectroscopy for the determination of ACs. Experimental considerations The basis of a reliable VCD-based assignment of an AC is high quality experimental IR and VCD data. While this means that the VCD spectroscopist needs to ensure a good signal to noise ratio, reduction of birefringence-induced baseline artefacts and baseline stability in general, the synthetic or natural products chemist worries about something else: How much sample do I need to provide? In this context, it generally needs to be stated that IR and VCD are rather insensitive methods and typically require concentrations in the range of 10-100 mM. In practice, this means that one measurement requires about 2-5 mg of sample, which can usually be recovered after the measurement. For synthetic samples and especially for those which are products of newly developed catalytic reactions, this amount is often easily prepared. Moreover, even if two to three runs are required to accumulate this amount, the time spent on repeating the synthesis several times (which is done in catalyst development anyways) is often shorter than the time spent on attempting to grow crystals suitable for crystallographic analysis. Typically, it is also not a problem to work with samples of low enantiomeric excess (e.e.) and 70% e.e. is often more than sufficient to determine the AC of the major isomer. Along with the required high concentrations often comes the problem of solubility. The most ideal solvents for VCD studies are chloroform-d1 (CDCl3) or carbon tetrachloride (CCl4), as they



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do not interact strongly with the sample. Acetonitrile-d3 (ACN-d3) and dimethylsulfoxide-d6 (DMSO-d6) are also often used and so are solvent mixtures. Common to these solvents is the fact that they do not absorb strongly in the mid-infrared region so that there is no (strong) overlap with bands of the solute. This is important as a VCD spectrum is a difference spectrum (VCD = A = ALCP-ARCP) and consequently, wherever band intensities are near or in total absorbance (typically: absorbance A > 0.9), it is not possible to reliably determine A. In turn, the solute concentration can also be too high, namely when the band intensities of solute vibrations themselves are above said threshold. Once the experimental conditions, i.e. concentration, solvent and path length, are optimized to an IR absorbance below 0.9 in the spectral range of interest, the VCD spectrum is recorded – depending on the signal intensity and thus the sample and its e.e. – within a few hours. The major work, though, is the computational analysis of the spectrum.

Theoretical prediction of IR and VCD spectra Besides recording high quality IR and VCD spectra, the computational analysis of the data is the key to the determination of an unknown AC. Today, this mostly follows a quite general protocol, which is outlined in Figure 1. Based on the chemical structure, which should at this point already be known from NMR spectroscopy and other common tools yielding information of atom connectivity, three dimensional computer models are generated for one enantiomer of each of the diastereomers which have to be considered (multiplication of the obtained VCD spectrum by -1 yields the spectrum of the corresponding opposite enantiomer). As all populated conformations of a molecule contribute to its IR and VCD spectra, the first key step of the computational workflow is a comprehensive conformational analysis. This is achieved by either systematically


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evaluating all possible combinations of dihedral angle settings, by statistical Monte-Carlo (MC) based searches or by the use of molecular dynamics (MD) simulations. Common to all methods for the initial conformer screening is the use of computationally inexpensive molecular mechanics force fields (FF). However, which of these methods is used depends mostly on the size of the molecule respectively on the number of degrees of conformational freedom. In the next step, starting with the lowest energy conformer of the FF-based search, a subset of structures is defined whose size is either determined by a certain energy threshold (e. g. 10 kcal/mol in force field energy) or on a simple number cut-off (e. g. the 200 lowest energy conformers). This subset is subject to further structural refinement on the density functional theory (DFT) level, usually employing common hybrid functionals such as B3LYP/B3PW91 (with or without dispersion corrections), a triple zeta basis set equipped with polarization and diffuse functions, and a continuum solvation model (e.g. the polarizable continuum model PCM20-21). However, in order to save computer time for large and complex molecules, the basis set size is occasionally reduced to double zeta. The geometry optimizations on DFT-level often significantly reduce the total number of individual conformers, as some of the FF-optimized structures converge to the same local DFT-minimum.



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Figure 1. Schematic workflow diagram for the theoretical prediction of VCD spectra.

For all remaining unique conformers, the relative energy differences and the vibrational spectra are computed. In order to determine the conformational distribution, i. e. the Boltzmann factors of all conformers, it is common to use either the Gibbs free energy G298K or the zeropoint corrected electronic energy EZPC. These populations are then used to compute the final IR and VCD spectra as the Boltzmann average of the single conformer spectra. Therefore, the computed stick spectra of each conformer, which consist of data point pairs of the frequency and either the dipole or the rotational strength for the IR and VCD spectra, are assigned a Lorentzian line shape (typically with a half-width at half height, HWHH, of 6 or 8 cm-1) to simulate the experimental band shapes. Afterwards, the Boltzmann factors are used to scale the contributions of each conformer. In addition to the line broadening, the frequency axis must usually be scaled


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by a factor  ranging from 1.02-0.96 in order to account for errors in the calculations of the vibrational frequencies due to the underlying harmonic approximation.

The comparison between experiment and theory Once both measurements and spectra simulations are completed, the comparison of the experimental and theoretical data should provide a very good match for one of the enantiomers, while the opposite enantiomer should show a mirror image relationship and hence hardly any match with the experiment. The initial assessment of the agreement is typically done visually by qualitatively comparing the experimental and theoretical spectral pattern. If there is no notable agreement or any doubt at this point, it is likely that the calculations can be improved by either checking for potentially missing conformations or changing the level of theory (e.g. increasing the basis set size or using a different functional). It may also be necessary to consider the environment, for instance by placing explicit solvent molecules near hydrogen bonding sites (microsolvation approach)7,

22

or by considering self-aggregation of the solute. How such

microsolvation can affect IR and VCD spectra has recently been discussed by us for the example of the carboxylic acid moiety.23-24 In a second step of comparison, the agreement is often quantified by the use of similarity analysis algorithms.25-26 There are different approaches for carrying out such analysis, but all rely on the calculation of the spectral overlap between the experimental and predicted spectral pattern. Among the most common, as they are implemented in free and commercially available software package,27-29 are the similarity measure  and the enantiomeric similarity index  (or ESI).27 Based on the overlap integral, the similarity measure  equals 1 for identical spectra and 1 when they are opposite to each other. For IR spectra, which do not have negative values, the



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lower boundary is 0 for spectra showing no similarity. The ESI complements the similarity analysis by simply representing the difference in the -values of the comparison between the experiment and either one of the enantiomers. The higher the value of  the better is the agreement with one of the possible enantiomers, while a low ESI indicates that the computed spectra of both enantiomers match equally well with the experimental (similar -values). The ESI is thus a powerful measure for the discriminating power of the predicted spectra. Recently, the vibrational dissymmetry factor (VDF), which is the ratio of VCD and IR spectrum (VDF=VCD/IR), has been introduced as another quantity for spectra comparison.25-26 The VDF has the advantage that it is concentration-independent, i.e. it can be determined even if the exact concentration of the sample is not known. Moreover, over- or underestimation of predicted band intensities is much better accounted for as well. The similarity measure  can also be used to quantify the agreement between experimental and theoretical VDF. A good match between experimental and predicted spectra is characterized by high similarity measures. Experimental factors such as noise but also slight differences in band position, width and intensity reduce the overlap integrals of the VCD spectra and practically do not allow values close to 1. Extensive evaluation of the approaches has thus led to the conclusion that VCD and  values below 0.5 and a VDF below 0.4 are not suitable to make an unequivocal assignment.29-31 Occasionally, VCD-based AC assignments also report a confidence level (CL). This measure has been introduced by Bultinck and co-workers and compares the VCD and of the molecule under consideration with datasets of about 80 AC determinations reported in literature.27 Based on this comparison, the CL algorithm is calculating percentage reliability for the assignment. As the database contains only very few misassigned structures, CL values below 95 % must be regarded with special caution. Moreover, unlike the other similarity measures, the CL is not suitable to


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distinguish different chemical structures as two diastereomers can both give high CLs.29Despite the availability of all the quantitative analysis tools introduced above, the visual agreement must be convincing and the similarity measures should be regarded as additional help to make an assignment and not its sole basis.

WORKED EXAMPLES FOR AC DETERMINATIONS Using three examples from our previous studies (cf. Scheme 2), this section demonstrates the above theoretical description of the AC determination procedure with real life examples.

Scheme 2. Structures of three molecules which will serve as models to demonstrate the above described AC determination procedure. The labelled stereocenters were determined by VCD spectroscopy.

Example 1: A single stereocenter in chiral cyclic lactones Certainly the easiest cases are those in which the VCD analysis has to differentiate between two stereoisomers of identical relative configuration, i.e. two enantiomers. In this case, as mentioned above, a calculated spectrum for an arbitrarily chosen enantiomer should either show a very good or almost no agreement with the experimental spectrum.



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Figure 2. Comparison of experimental IR, vibrational CD and vibrational dissymmetry spectra (black/grey traces) with computed spectra (red) of (S)-1. Experimental conditions: 0.335 M, CDCl3, 100 µm path length; Computational parameters: B3LYP/6-311+G(2d,p) with IEFPCM(CHCl3), 8 cm-1 HWHH; Similarity analysis: =0.99, IR=91% , VCD=76 %, =72 %, VDF=42 %

The first worked example is taken from a study on biocatalytic redoxisomerizations in which an alcohol dehydrogenase was used to produce various γ-hydroxy-δ-lactones in high enantiopurity.32 We herein focus on 4-hydroxyisochroman-1-one 1, whose experimental IR and VCD spectra were recorded for a 0.335 M solution in chloroform-d1 (Figure 2). Due to the small size of the molecule, the conformational analysis was carried out systematically by considering the rotation of the OH-group and two conformations of the lactone ring. The three lowest energy conformations of (S)-1 and their relative Gibbs free energies G298K and Boltzmann populations are shown in Figure 3. Based on the single conformer spectra, the simulated spectra shown in


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Figure 2 were obtained. Already on first sight, the simulated spectra provide a striking match with the experimentally observed IR and VCD spectral pattern. Solely the predicted positive feature at 1100 cm-1 is not found in the experiment. Based on this visual comparison, the configuration can thus unambiguously be assigned to be (S), as an (R)-configuration would require the experimental spectrum to match with a mirrored (signs inverted) simulated VCD spectrum. A quantitative comparison using the above mentioned similarity measures further supports this assignment: VCD = 76 % and = 72%.

Figure 3. The three lowest energy conformations of (S)-4-hydroxyisochroman-1-one 1 and their relative Gibbs free energies G298K and Boltzmann populations p obtained at B3LYP/6311+G(2d,p) with IEFPCM(CHCl3)

In addition to the IR and VCD spectra, Figure 2 also shows the experimental and simulated VDF spectrum. Typically, in order to avoid large contributions of noise and thus to facilitate quantitative comparison, the experimental VDF spectrum is set to zero in spectral ranges where there are no IR bands observable (i.e. where the molar absorptivity is below a certain threshold). In case of 1, this is, for instance, done in the range from approximately 1580-1500 cm-1. While the qualitatively good match is obvious, the similarity of VDF spectraVDF can be calculated to further increase the confidence by using the same algorithm which is used to calculate VCD and . For the VDF spectra in Figure 2, a rather small value of VDF = 42 % is obtained, most likely due to the intense signal at around 1350 cm-1. In this range, the corresponding IR intensities are



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slightly above the zeroing-threshold, hence causing a large noise signal. Nevertheless, together with be VCD and  the assignment is confirmed without any doubt.

Worked examples 2 and 3: Differentiating diastereomers Diastereomers can often already be distinguished based on a detailed NMR spectroscopic analysis by taking advantage of the torsional angle dependence of coupling constants or 1H,1HNOE contacts.33 Once the relative configuration is completely known, it is only necessary to distinguish two enantiomers and hence the determination of the AC using VCD spectroscopy can proceed as outlined. It becomes more challenging, when relative configurations cannot be established due to missing 1H,1H-interactions, for instance because one of the stereocenters is a quaternary carbon center (such as in 2), or because of an inconclusive coupling pattern (as found for 3). In these cases it is necessary to compute the spectra for all possible diastereomers (respectively one enantiomer of each) and carefully evaluate the agreement with the experimental spectra. The example of 2 (cf. Scheme 2) is taken from a recent study on the development of a new asymmetric catalyst.34 Due to detailed knowledge about the reaction mechanism of the catalyst used to prepare the compound, the bridge head configurations and quaternary stereocenter were known, but the tertiary stereocenter could not be characterized. Therefore, models for both the (S,S)- and the (S,R)-configuration were generated and subjected to a systematic conformational analysis. As the general scaffold of 2 is fairly rigid, the conformational analysis revealed only two to three dominating conformations.


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Figure 4. Comparison of the experimental IR, VCD and VDF spectra of 2 with the predicted spectra of (S,S)- and (S,R)-2. Experimental conditions: 87 mM, CDCl3, 100 µm path length; Computational parameters: B3LYP/6-31+G(2d,p) with IEFPCM(CHCl3), 6 cm-1 HWHH. cf. Table 1 for similarity analysis.

Figure 4 shows the computed spectra of (S,S)- and (S,R)-2 in comparison with the experimental spectra, which were recorded for an 87 mM solution of 2 in chloroform-d1. Visual comparison of the predicted IR spectra reveals only small differences, with the most important ones being notable in the carbonyl stretching region (1800-1650 cm-1). Especially in this region, the computed spectrum for (S,S)-2 agrees qualitatively better with the experiment, which is, however, not sufficient for an unambiguous assignment. The predicted VCD spectra reveal several bands, which appear to be characteristic for the respective diastereomer. In Figure 4, several of these characteristic bands are marked with (+) and (-) signs, which are meant to help



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the reader to perform a simple pattern comparison and qualitatively conclude the assignment of an (S,S)-configuration. Despite some experimental noise in the range of 1400-1300 cm-1, the comparison of the VDF spectra further supports this assignment. Lastly, the quantitative similarity analysis provided in Table 1 gives the final confirmation of the assignment, as the similarity between the experimental spectra and those calculated for (S,S)-2 are clearly larger than for (S,R)-2.

Table 1. Quantitative comparison of the experimental spectra of 2 with the two considered diastereomers (S,S)- and (S,R)-2. calcd.



IR

VCD



VDF

(S,S)

0.98

83

63

58

60

(S,R)

0.985

84

24

2

10

Occasionally, VCD spectroscopy is required to distinguish two diastereomers without previous knowledge about the complete relative configuration of the experimentally available samples. There is, however, no guarantee that the differences between the spectra of the diastereomers are as pronounced as they are in the case of 2. Therefore, it may be advantageous to have both diastereomers available for measurements. We exemplify the problem for the example of the diastereomers of 3 (cf. Scheme 2), synthetic substrate surrogates used in a recent study on the ambruticin biosynthesis.35 All stereocenters indicated in the structure shown in Scheme 2 were known from previous studies, and the unknown configuration at the -position of the alkyl chain was not accessible via NMR spectroscopy due to too high similarity of coupling constants. As the side chain features a high conformational flexibility, a Monte Carlo search algorithm was used in the initial conformational search on a FF level. For both diastereomers, this search generated thousands of conformers, of which the 100 lowest energy conformers of each of the


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sets were subjected to further geometry optimizations as well as IR and VCD spectra calculations. Due to the large number of structures, we lowered the basis set size and used the B3LYP/6-31G(2d,p)/IEFPCM(CHCl3) level of theory for all calculations. For the simulation of the final Boltzmann weighted IR and VCD spectra, contributions of merely 14-18 conformers had to be considered.

Figure 5. Comparison of the experimental and computed IR and vibrational CD spectra of dia13 and dia2-3 as well as of the difference spectrum [dia2]-[dia1] and its computational counterpart. The (+) and (-) signs indicate the characteristic differences between the diastereomers’ spectra. Experimental conditions: 0.2 M, CDCl3, 100 µm path length; Computational parameters: B3LYP/6-31G(2d,p) with IEFPCM(CHCl3), 6 cm-1 HWHH; cf. Table 2 for similarity analysis. Figure 5 presents the experimental and simulated IR and VCD spectra of the two diastereomers of 3 in a slightly different way than in the figures discussed above. Instead of stacking the spectra and showing band correlations as in Figures 2 and 4, we separately overlay the sets of



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experimental and theoretical spectra in order to emphasize the differences between them. In the range from 1450-1350 cm-1 and even more obviously in the range below 1150 cm-1, these small differences between the experimental VCD spectral patterns can be noted. Besides a general band shift, the same small differences can be recognized in the theoretical spectra. Differences in the IR spectra are less pronounced, but also clearly visible. Hence, an assignment of diastereomer 1 to the (R)- and diastereomer 2 to the (S)-configuration can be qualitatively established.

Table 2. Similarity analysis of dia1- and dia2-3 measured over the range 1600-950 cm-1. exptl

calcd



IR

VCD



VDF

Dia1

(R)

0.98

95

67

64

51

(S)

0.98

92

26

7

6

(R)

0.98

93

69

63

47

(aS)

0.98

94

46

26

43

(S)-(R)

0.985

47(a)

36(a)

Dia2

Dia2-Dia1

(a) Calculated for the difference spectra

A trained user can thus make a reliable assignment by visual comparison, potentially even if only one isomer was available. However, as shown in Table 3, the similarity analysis algorithms have a much harder time in differentiating these stereoisomers. In fact, when compared with each set of experimental spectra, the (R)-isomer yields a higher similarity than the (S)-isomer in both cases. We attribute this to the overestimation of the predicted intensities in the spectral range below 1150 cm-1, which have a large weight in the overlap analysis. As both diastereomers are available, a well-established strategy can be applied which is based on the difference of the experimental theoretical spectra, respectively:36 it compares the calculated difference spectra


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[dia2]-[dia1] and [S]-[R]. From comparison of these difference spectra, both qualitatively and quantitatively, the stereochemical relationship becomes immediately obvious (Fig. 5, top; Table 2).

DEMONSTRATING THE DIVERSITY OF AC DETERMINATIONS CARRIED OUT USING VCD SPECTROSCOPY The worked examples presented in the previous section outlined general strategies in analysing IR and VCD spectra for the determination of ACs. Within this section, we want to provide a review of VCD-based AC determinations carried out in the past decade (period 2008-2018) in order to showcase the structural variety of molecules characterizable by VCD spectroscopy. We selected only AC determinations of fully synthetic compounds, for which one or more stereocenters were generated during the synthesis. In other words, we intentionally left out studies on molecules whose ACs were unaffected by the reaction. Moreover, studies on natural products and their derivatives were left out, as there is a very comprehensive recent review by Batista et al.1 It should nevertheless be noted that the herein targeted AC determinations are often not in the focus of the main articles and thus buried in supporting information files. We will therefore not claim completeness of this review section, although we have attempted to be as thorough as possible.

Tertiary stereocenters Enantioselective catalysis gives access to enantiopure or at least significantly enantio-enriched products. However, especially when yet unprecedented transformations with known or new asymmetric catalysts are attempted, the stereochemistry of the chiral products must often still be



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determined in order to either confirm the stereoselectivity or to understand the mechanism of stereoinduction. The vast majority of asymmetric catalytic reactions targets the generation of tertiary stereocenters, and often only one stereocenter per molecule is generated during the course of these reactions. Moreover, the target molecules often carry a variety of functional groups giving rise to rich IR and VCD spectra. Therefore, VCD spectroscopy is often employed to assign the AC of the tertiary carbon centers by basically following a similar route as discussed for example 1 above (cf. Table 3). Examples for such reactions and applications of VCD are manifold and can be found for both transition metal (TM) as well as organocatalytic reactions. Lacour and co-workers, for instance, reported the regioselective and enantiospecific synthesis of dioxepines like 4 by condensation of diazocarbonyls and chiral oxetanes.37 In order to rationalize the mechanism of the reaction, which involved a (cyclopentadienyl)ruthenium complex and 1,10‐phenanthroline, it was necessary to confirm the AC of the product. Using VCD spectroscopy, it was shown that the initial configuration of chiral oxetane starting material is retained in the final dioxepines. For 5, the group followed a similar strategy.38 Likewise, DFT-based predictions of the stereochemistry of a stereoselective copper-bis(oxazoline)-catalyzed carbene insertion reaction between methyl diazophenylacetate and tetrahydrofuran, which gave access to enantio-enriched 6 and derivatives thereof, was confirmed by VCD spectroscopy.39 In context of the confirmation of the stereochemistry of products of TM-catalyzed reactions, further studies are worth noting. There are, for instance, 7 and its derivatives, which are the products of an asymmetric C(sp3)-H/C(Ar) coupling reaction of indoline catalyzed by a chiral NHC-palladium complex.40 The AC of 8 was reported in a study on the Rh- or Ir-catalyzed


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directed C(sp3)–H borylation, in which Reyes et al. used chiral phosphoramidite ligands in order to induce enantioselectivity to the reaction pathways.41 Likewise, the enantioselective addition of boronic acids to N-benzylnicotinate salts was achieved via Rh(COD)2BF4/(S)-BINAP catalysis yielding 9.42 Table 3. AC determination of molecules with tertiary stereocenters

7

4

5

6

8

9

10

11

12

13

14

15

16

17

18

19

21

22

23

25

26

27

20

24

Newly developed organocatalytic reactions benefit similarly from unambiguous AC determinations using VCD spectroscopy. It has for instance been applied of in the characterization of chiral 3-substituted isoindolinones like 10, which were obtained using a thiourea derived from quinine.43 Likewise, the stereochemistry of the tertiary stereocenter of 11,



Page 22 of 50

which were obtained through an asymmetric epoxidation of 2‐arylidene‐1,3‐diketones, was determined in dependently by OR and VCD spectroscopy.44 Stereocenters cannot always be created through enantioselective reactions and optical resolution by diastereomeric crystallization or chiral HPLC is often required to obtain enantiopure compounds. Among these are several biologically and pharmacologically active and thus relevant compounds. Lactofen 1245, for instance, is an herbicide used to control broadleaved weeds in certain crop fields, that is typically applied in its racemic form. In order to investigate the different herbicidal activities of the enantiomers of 12, they were separated by chiral HPLC and characterized by VCD and ECD. 13 is a potential antagonist for the transient receptor potential melastatin type 8 (TRPM8), a nonselective cation channel expressed in sensory neurons that is under investigate for new approach to treat pain.46 14 is one of three potential A3 adenosine receptor antagonists investigated by Rossi et al., which may be of interest as cardioprotective agents, as anti-inflammatory and immunosuppressive agents, as cytostatics and chemo-protective compounds in cancer therapy.47 15 is an inhibitor for sirtuins (SIRTs), an enzyme involved in aging-related diseases.48 Further examples are: an intermediate in the synthesis of an phthalazine-based p38 MAP kinase inhibitor (16),49 the aroma compound 2,5-dimethyl-4-methoxy-3(2H)-furanone 17 and several of its derivatives;50 the 2,6-bis(2-butyl)-4-methyl-pyridine 18 and its N-oxide;51 aromatase-sulfatase inhibitor 19, for which no suitable crystals could be obtained;52 the α(trifluoromethyl)-tryptamine 20;53 the chiral sulfonamide 21;54 a new calcium channel blocker 2255; products of the newly developed biocatalytic reduction of the corresponding imines to sulfur containing cyclic amines such as 2356; the monoamine oxidase inhibitor 2457; the


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phenothiazine-based derivative 25,58 the acetonide 26,59 the dihydronaphthalene 27,60 which is chiral by isotopic labelling only. Table 4. AC determination of molecules with quarternary stereocenters

31 28

29

30

NO2 Me O Me

32

33 34

35

39 36

37

38

40

40

41

One quarternary stereo center In enantioselective synthesis, the formation of quarternary carbons as stereocenters is of particular interest but in many cases the help of VCD spectroscopy is required to determine the ACs (cf. Table 4 for examples). The C2-stereocenter of dihydrofuran 28, for instance, was introduced by a palladium-catalyzed asymmetric Heck reaction of dihydrofurans with a trisubstituted double bond.61 In this reaction, the choice of the phosphine ligands was found to determine whether 2,3- or 2,5-dihydrofurans are formed. As no suitable crystals for x-ray crystallography could be obtained, VCD has been used for the AC determination. For the preparation of 29, a precursor to hinckdentine A, a natural product that can be isolated from the marine bryozoan Hincksinoflustra denticulate, Douki et al. also utilized a Heck type reaction.62 They used both ECD and VCD to unambiguously assign its AC. This provided increased confidence in



Page 24 of 50

the assignment by the combined use of two techniques, but for the given example one of them would have sufficed. Cyclic carbonates and oxazolidinones such as 30 were obtained from the reaction of CO2 with propargylic alcohols respectively propargylic amines through a [3,3]sigmatropic rearrangement in the presence of AgOAc and a chiral Schiff base.63 Also in the presence of a silver salt, silver phosphate, and catalytic amounts of Pd(OAc)2 and chiral bis(oxazoline) ligands, the spirolactam 31 was obtained by an carbocyclization cascade during which all stereocenters were generated simultaneously in high diastereomeric ratio (dr, >99:1).64 While the AC of 31 was determined solely based on VCD spectroscopic data, also x-ray crystallographic data was available for 30.63 Examples for AC determinations of quarternary stereocenters can also be found in the field of asymmetric organocatalysis. Wang et al. reported the use of Takemoto's catalyst65 for an intermolecular asymmetric dearomatization reaction, which transformed β‐naphthols with nitroethylene through a Michael reaction to structures like 32 with an all-quaternary stereocenter.66 The all‐carbon quaternary β‐stereocenter of γ‐butyrolactones of the type of 32 were enantioselectively prepared by a tandem aldol/lactonization sequence in presence of a cinchona alkaloid-derived organocatalyst.67 Further examples are: propellane 34 (which may be considered axially chiral as well);68 the building block for αvβ6 integrin inhibitors 35;69 the products of the anodic oxidation of 2,4dimethylphenol such as 36;70 the bioactive spiropyrazolone derivatives 37;71 the lactone 38.72 Stoltz and co-workers investigated the enantioselective Pd-catalyszed allylic alkylation yielding dialkylated N-heterocycles such as 39 and utilized VCD spectroscopy to determine the AC of several of the obtained products.73


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Compounds 40, 41 and 4274 are interesting examples from a field of asymmetric synthesis, which takes advantage of the concept of memory of chirality (MOC).75 A particular characteristic of such MOC reactions is that they take place via chiral enolate or radical species with rather short half-life times of racemization which leads to a competition between the actual reaction of the chiral intermediate with its racemization. The benzocyclobutenone 40,76 for instance, is the product of an asymmetric -aryllation of protected -amino acids which proceeded via an intermediate enolate. Azaheterocylic 41 and further derivatives with a quarternary stereocenter were obtained from the cascade rearrangement of chiral enediynes, which proceeded through radical intermediates.77

More than one stereogenic center VCD spectroscopy shows its full power when more than one stereocenter has to be determined simultaneously. With 7 and 31, two such cases have been shown already and Table 5 summarizes further examples. In fact, asymmetric reactions often yield diastereomerically pure products as several stereocenters are generated simultaneously and not completely independent of each other. A typical example for such reactions is the enantioselective epoxidation of asymmetrically substituted olefins, which leads from -ylideneoxindoles leading to structures like to 43.78 Similarly, asymmetric Sharpless dihydroxylations lead to diols like 44.79 In many such cases, the relative configuration can often easily be established based on NMR data (utilizing either scalar or through space coupling), so that VCD spectroscopy is only required to establish the AC.



Page 26 of 50

Table 5. AC determinations of diastereomers

46 43

47

44 45

52 48

49

50

51

53

54

55

56

58

59

60

61

57

Diels Alder reactions and reactions on bicyclic system are also often found to proceed with high diastereoselectivity. Starting from norcamphor, Lattanzi et al. prepared 45 and determined the AC of the C2-stereocenter independently by OR and VCD spectroscopy and the corresponding calculations.80 The endo- and exo-forms both showed positive OR values, so that a differentiation was only possible based on a comparison of the relative size of the computed values. The VCD spectra of the diastereomers, which are reproduced in Figure 6, were found to feature characteristic marker bands in the spectral range from 1200-1000 cm-1, so that the isomers could easily be distinguished solely based on their VCD signatures. The trifluoromethyl‐ substituted diarylprolinol silyl ether catalysed Diels Alder reaction between acrolein derivatives and cyclopentadiene, for instance, yielded the exo-products in excellent dr and good e.e. A


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subsequent reduction and treatment with I2 yielded iodolactone 46 and an iodoether, which were easily separated by column chromatography.81

Figure 6. Comparison of the experimental VCD spectrum of 45 recorded in CCl4 with the computed spectra of the exo- and endo-form as obtained at the B3PW91/TZVP level of theory. The presented data was digitalized from Ref.

80

and the frequency axis of the computed spectra

was scaled by =0.975.

An impressive example for a highly diastereoselective reaction is the aza-Michael-initiated ring-closure reaction of nosyloxycarbamates. Unexpectedly, the investigated ring-closure reaction was found to proceed stereospecifically, as only one out of six pairs of stereoisomers was produced and identified by NMR spectroscopy. After separation of the enantiomers on a chiral HPLC, the AC of the chiral 2,2'-dinitro-2,2'-biaziridine 47 has been determined using ECD and VCD spectroscopy.82 Similarly, the AC determination of 48 helped in elucidating the mechanism of a diarylprolinol silyl ether catalysed intramolecular [6+2] cycloaddition.83 Via an unexpected rearrangement reaction, Goijer et al. obtained the 3,7-diasteroemers of 49 from the reaction of enantiopure 3-aminequinuclidine and 2-chloropyrimidine.84 Again, the



Page 28 of 50

diastereomers could be differentiated based on NMR spectroscopic, and the AC was determined by VCD after HPLC separations of the diastereomers. Further examples for diastereomers, which were characterized by VCD spectroscopy are: the β-fluoro cyclobutylimine 50,85 the unusual anti aryl β-hydroxy α-amino ester 51,86 the aldolproduct 52,87 the potential KDM1A inhibitor 53,88 the menthone derivative 54;89 the decahydro4,8-epoxyazulene 5590 and troponoids 56,72 both products of Rh(II) catalyzed enantioselective reactions; the tricyclic product 57,91 which is obtained via a catalytic aerobic oxidation and tandem enantioselective cycloaddition in a cascade multicomponent synthesis. Despite usually being quite reliable, there are also cases in which the classical tools used to determine relative configurations do not give clear answers. In case of 58, for instance, not only the quaternary stereocenter but also the relative configuration of the other substituents had to be confirmed by VCD spectroscopy, as the computational analysis of the reaction mechanism and crystallography data were not conclusive enough.92 Somewhat related are examples like brivaracetam 59, for which the AC had initially been determined based on the known stereochemistry of the starting material and an x-ray structure which gave access to the relative configuration. With VCD, both stereocenters of 59 could be determined simultaneously. Covalently bound chiral auxiliaries are often employed in order to induce stereochemical preferences during the course of a reaction or in order to obtain easily separable diastereomers. The molecules 60 and 61 are examples for such diastereomers which were prepared with menthol93 and -phenylethyl amine94 as auxiliaries, separated by chromatography and lastly analysed by VCD spectroscopy.

Aliphatic molecules and cryptochirality


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Purely aliphatic compounds impose a particular challenge for AC determinations as they carry no suitable UV/Vis chromophores required to record ECD spectra (cf. Table 6). A good example for such a case is molecule 62, which has been prepared by Maes and co-workers through a novel Ru-catalyzed C(sp3)-H functionalization.95 Somewhat similar is the situation for the chiral cyclohexanone 63,96 which was obtained by an asymmetric copper-catalysed alkylation reaction. Here the stereocenter on the flexible alkyl chain imposes a particular challenge for ECD spectroscopy, as there is no UV-active chromophore in its vicinity and the carbonyl group cannot serve as probe for both stereocenters at the same time. While 62 and 63 still have UV absorbing chromophores, cryptochiral compounds do not. Hence, they also feature barely any optical rotation. Three such examples were characterized by VCD spectroscopy, namely d6-neopentane 6497, enantiopure 4-ethyl-4-methyloctane 6598 or the -helicene 66.99 As an example, Figure 7 shows the VCD spectra of 65, in which the spectral signatures arise only from C-C stretching and CHx-bending modes. In contrast, while the VCD spectrum obviously features several bands that allow an unambiguous assignment of the AC, the optical rotation measured at 365 nm was only -0.73° for the neat liquid and hence did not allow any reliable assignments. Table 6. AC determination of aliphatic and cryptochiral molecules

64 62

65

66

63



Page 30 of 50

Figure 7. Comparison of the experimental and the calculated VCD spectrum of the colorless oil (S)-(-)-65 ([α]36521 = –0.73 for the neat liquid with a density of ρ = 0.7565). The data was digitally extracted from Ref.

98

The original spectra calculations were performed on the

B3PW91/6-31G(d,p) level of theory and a line broadening of 8 cm-1 HWHH was applied. For the presentation herein, the frequency axis of the computed spectrum was scaled by =0.98.

Axially and planar chiral molecules and other topologies The examples mentioned up to this point mostly featured tetrahedral carbon atoms as centers of chirality. VCD is of course not limited to central chirality and can also be applied to other types of chiral molecules (cf. Table 7 for examples). Allenes are the simplest class of axially chiral molecules. They can be obtained from suitable prochiral precursors such as prochiral dichloro allenes. By treatment with Grignard reagents in the presence of copper salt and a chiral ligand, an asymmetric propargylic substitution gave access to allenes with quaternary stereogenic centers such as 67.100 Propellanes such as 34 may also be considered axially chiral.68 Generally, however, the molecular dimensions must be larger in order to establish axial chirality. In 68, for instance, the rotation along the aryl-aryl bond is hindered due to the bridging side arms, so that racemization is efficiently prevented.101 In 69, the stereochemistry of the rigid binapthyl-unit induces a conformational preference in the bipyridine unit.102 In the axially chiral 70,103 axial


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chirality is established in a bis(dinaphthofuran)moiety. There does not have to be a linking bridge as shown in the examples of the 4‐arylisoquinolone 71104 and nitroxide 72.105 In both cases, the substitution pattern around the axis provides a sufficiently high energy barrier to prevent racemization. Likewise, in 73, the chiral side arms induce axial chirality to the 3,3’bithiophene unit, which is even preserved after removal of the chiral oxazoline moieties.106 Similar concepts were realized in the examples 74107 and 75108 in which methyl substitutions are sufficient to prevent isomerization. In 76, the sulfonate acts as bulky configurational anchor and hinders the rotation of the naphthyl groups.109 Table 7. Examples for axially and planar chiral molecules characterized by VCD spectroscopy.

71 67

68

69

70

72

73

74

75

76

77

78

79

80

81

82

83

84

85

The bowl shaped molecules 77110 and 78111 show that there are many other ways to realize axial chirality. Helicenes (79)112-113 can be considered as the prototype molecules for axial chirality and



Page 32 of 50

they usually feature characteristic signatures in electronic CD spectra due to their extended system. Especially the group of Crassous has carried out plenty of VCD studies on various derivatives and metal complexes114-115 as well as helicene-like molecules116 such as 80.117 Further examples are atropoisomers of the 10-membered ring with multiple chirality axes 81,118, the axially chiral carbodiimide 82119 or the atropisomers of the EE-tetrahalogeno-1,3-butadiene core 83.120 Cyclophanes such as 84 are classical examples for planar chirality.121-122 Also 85, a product of a [2+2+2]-cycloaddition reaction, can be considered a cyclophane and must be classified as planar chiral as it does not feature any stereocenter or stereoaxis that could be used for a definition.123

Non-carbon stereocenters The electron lone pair serves as fourth substituent on the tetrahedral sulfur atom of sulfoxides, so that a sufficiently high energy barrier for the sulfur inversion allows the efficient stabilization of either of the enantiomers (86-88, Table 8). These can be synthesized by enantioselective oxidation of the corresponding sulfide, as it has been shown for 86,124 prepared along a diastereomeric synthesis route as in the case of phenyl perdeuteriophenyl sulfoxide 87,125 or simply resolved by chiral HPLC as done for 88.126 87 is particularly noteworthy as it is the first isotopically chiral sulfoxide analysed by VCD spectroscopy. The phosphine-borane complex 89 features a tetrahedral phosphorous atom with four different substituents. Enantiopure 89 was prepared by resolution of the racemate on chiral HPLC and the AC was independently determined by x-ray crystallography and VCD.127 Deprotection even yielded solution-stable enantiopure phosphine. Pentacoordinate chiral phosphorus compounds 90 were prepared from enantiopure amino acids,128 whereby the phosphorus atom became center of


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axial chirality. The determination of the configuration at the pentacoordinate center was achieved by VCD spectroscopy, and it was found that the carbonyl stretching region is particularly sensitive for the phosphorus chirality due to exciton coupling of the carbonyl modes,129 while the remaining spectral range (

Absolute configurations of synthetic molecular scaffolds from

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