Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Protocol
  • Published:

Interrogating the configurational stability of atropisomers

Nature Protocols volume 18, pages 2745–2771 (2023)Cite this article

Subjects

Abstract

Atropisomers are molecules whose stereogenicity arises from restricted rotation about a single bond. They are of current importance because of their applications in catalysis, medicine and materials science. The defining feature of atropisomeric molecules is that their stereoisomers are related to one another by bond rotation: as a result, evaluating their configurational stability (i.e., the rate at which their stereoisomers interconvert) is central to any work in this area. Important atropisomeric scaffolds include C–C linked biaryls, such as the ligand BINAP and the drug vancomycin, and C–N linked amine derivatives such as the drug telenzepine. This article focuses on the three most widely used experimental methods that are available to measure the rate of racemization in atropisomers, namely: (i) kinetic analysis of the racemization of an enantioenriched sample, (ii) dynamic HPLC and (iii) variable-temperature NMR. For each technique, an explanation of the theory is set out, followed by a detailed experimental procedure. A discussion is also included of which technique to try when confronted with a new molecular structure whose properties are not yet known. None of the three procedures require complex experimental techniques, and all can be performed by using standard analytical equipment (NMR and HPLC). The time taken to determine a racemization rate depends on which experimental method is required, but for a new compound it is generally possible to measure a racemization rate in <1 d.

Key points

  • Stereoisomers of atropisomeric molecules interconvert by rotation of a single bond. If the interconversion rate is slow enough, the atropisomers can be separated by HPLC. After enrichment of one isomer, the kinetics of racemization can be determined.

  • At increasing rates of interconversion, analytical HPLC shows two peaks, a ‘Batman’ profile (dynamic HPLC) or a single peak.

  • For molecules with faster interconversion, variable-temperature NMR can be performed.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: An overview of atropisomerism and configurational stability.
Fig. 2: Different timescales for bond rotation and definitions of atropisomerism.
Fig. 3: Kinetic analysis of the racemization of an enantioenriched sample.
Fig. 4: Representative example.
Fig. 5: Dynamic HPLC and calculations using the unified equation.
Fig. 6: Representative example.
Fig. 7: Schematic example of an axially chiral biaryl.
Fig. 8: Representative example.
Fig. 9: Workplan for analyzing the barrier of a completely unknown compound.
Fig. 10: Analytical HPLC data for illustrative compound 1.
Fig. 11: Semi-preparative HPLC isolation of a single enantiomer.
Fig. 12: Monitoring atropisomer racemization by HPLC at elevated temperature.
Fig. 13: Using ΔG‡ to estimate racemization rates at other temperatures by using the ‘Eyring equation calculator’.
Fig. 14: Analytical HPLC data for illustrative compound rac-2.
Fig. 15: Analytical HPLC data for illustrative compound rac-2.
Fig. 16: 1H NMR spectrum of 4 at 20 °C in d8-toluene.
Fig. 17: 1H VT-NMR spectra of 4 in d8-toluene.
Fig. 18: DNMR in TopSpin.

Similar content being viewed by others

Data availability

We have previously reported synthetic procedures and data for compounds 1 (ref. 18), 2 (ref. 11) and 4 (ref. 11), which are used to illustrate all three procedures.

References

  1. Oki, M. Topics in Stereochemistry. Vol. 1 Atropisomerism (Wiley Interscience, 1983).

  2. Noyori, R. Asymmetric catalysis: science and opportunities (Nobel Lecture). Angew. Chem. Int. Ed. Engl. 41, 2008–2022 (2002).

    Article  CAS  PubMed  Google Scholar 

  3. Wencel-Delord, J., Panossian, A., Leroux, F. R. & Colobert, F. Recent advances and new concepts for the synthesis of axially stereoenriched biaryls. Chem. Soc. Rev. 44, 3418–3430 (2015).

    Article  CAS  PubMed  Google Scholar 

  4. Cheng, J. K., Xiang, S.-H., Li, S., Ye, L. & Tan, B. Recent advances in catalytic asymmetric construction of atropisomers. Chem. Rev. 121, 4805–4902 (2021).

    Article  CAS  PubMed  Google Scholar 

  5. Sweet, J. S. & Knipe, P. C. Catalytic enantioselective synthesis of C–N atropisomeric heterobiaryls. Synthesis 54, 2119–2132 (2022).

    Article  CAS  Google Scholar 

  6. Kumarasamy, E., Raghunathan, R., Sibi, M. P. & Sivaguru, J. Nonbiaryl and heterobiaryl atropisomers: molecular templates with promise for atropselective chemical transformations. Chem. Rev. 115, 11239–11300 (2015).

    Article  CAS  PubMed  Google Scholar 

  7. Mei, G.-J., Koay, W. L., Guan, C.-Y. & Lu, Y. Atropisomers beyond the C–C axial chirality: advances in catalytic asymmetric synthesis. Chem 8, 1855–1893 (2022).

    Article  CAS  Google Scholar 

  8. Rodríguez-Salamanca, P., Fernández, R., Hornillos, V. & Lassaletta, J. M. Asymmetric synthesis of axially chiral C−N atropisomers. Chemistry 28, e202104442 (2022).

    Article  PubMed  PubMed Central  Google Scholar 

  9. Wu, Y.-J., Liao, G. & Shi, B.-F. Stereoselective construction of atropisomers featuring a C–N chiral axis. Green. Synth. Catal. 3, 117–136 (2022).

    Article  Google Scholar 

  10. Clayden, J. Atropisomers and near-atropisomers: achieving stereoselectivity by exploiting the conformational preferences of aromatic amides. Chem. Commun. (Camb.) 2004, 127–135 (2004).

    Article  Google Scholar 

  11. Costil, R., Sterling, A. J., Duarte, F. & Clayden, J. Atropisomerism in diarylamines: structural requirements and mechanisms of conformational interconversion. Angew. Chem. Int. Ed. Engl. 59, 18670–18678 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Vaidya, S. D., Toenjes, S. T., Yamamoto, N., Maddox, S. M. & Gustafson, J. L. Catalytic atroposelective synthesis of N-aryl quinoid compounds. J. Am. Chem. Soc. 142, 2198–2203 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Toenjes, S. T. & Gustafson, J. L. Atropisomerism in medicinal chemistry: challenges and opportunities. Future Med. Chem. 10, 409–422 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Rickhaus, M., Jundt, L. & Mayor, M. Determining inversion barriers in atropisomers—a tutorial for organic chemists. Chimia 70, 192–192 (2016).

    Article  CAS  PubMed  Google Scholar 

  15. Reist, M., Testa, B., Carrupt, P.-A., Jung, M. & Schurig, V. Racemization, enantiomerization, diastereomerization, and epimerization: their meaning and pharmacological significance. Chirality 7, 396–400 (1995).

    Article  CAS  Google Scholar 

  16. LaPlante, S. R., Edwards, P. J., Fader, L. D., Jakalian, A. & Hucke, O. Revealing atropisomer axial chirality in drug discovery. ChemMedChem 6, 505–513 (2011).

    Article  CAS  PubMed  Google Scholar 

  17. Bragg, R. A., Clayden, J., Morris, G. A. & Pink, J. H. Stereodynamics of bond rotation in tertiary aromatic amides. Chemistry 8, 1279–1289 (2002).

    Article  CAS  PubMed  Google Scholar 

  18. Jolliffe, J. D., Armstrong, R. J. & Smith, M. D. Catalytic enantioselective synthesis of atropisomeric biaryls by a cation-directed O-alkylation. Nat. Chem. 9, 558–562 (2017).

    Article  CAS  PubMed  Google Scholar 

  19. Trapp, O., Schoetz, G. & Schurig, V. Determination of enantiomerization barriers by dynamic and stopped-flow chromatographic methods. Chirality 13, 403–414 (2001).

    Article  CAS  PubMed  Google Scholar 

  20. Trapp, O. Unified equation for access to rate constants of first-order reactions in dynamic and on-column reaction chromatography. Anal. Chem. 78, 189–198 (2006).

    Article  CAS  PubMed  Google Scholar 

  21. Trapp, O. Fast and precise access to enantiomerization rate constants in dynamic chromatography. Chirality 18, 489–497 (2006).

    Article  CAS  PubMed  Google Scholar 

  22. Trapp, O. The unified equation for the evaluation of degenerated first-order reactions in dynamic electrophoresis. Electrophoresis 27, 2999–3006 (2006).

    Article  CAS  PubMed  Google Scholar 

  23. Trapp, O. Interconversion of stereochemically labile enantiomers (enantiomerization). Top. Curr. Chem. 341, 231–270 (2013).

    Article  CAS  PubMed  Google Scholar 

  24. DCXplorer MCXVII download link: https://www.cup.lmu.de/oc/trapp/tools.html (2023).

  25. Lanman, B. A., Parsons, A. T. & Zech, S. G. Addressing atropisomerism in the development of sotorasib, a covalent inhibitor of KRAS G12C: structural, analytical, and synthetic considerations. Acc. Chem. Res. 55, 2892–2903 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Hirsch, D. R. et al. Troponoid atropisomerism: studies on the configurational stability of tropone-amide chiral axes. Org. Lett. 21, 2412–2415 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Clark, A. J. et al. Axially chiral enamides: substituent effects, rotation barriers, and implications for their cyclization reactions. J. Org. Chem. 81, 5547–5565 (2016).

    Article  CAS  PubMed  Google Scholar 

  28. Claridge, T. D. W. Introducing high-resolution NMR. In High-Resolution NMR Techniques in Organic Chemistry Edn. 3 (Elsevier, 2016).

  29. Kost, D., Carlson, E. H. & Raban, M. The validity of approximate equations for kc in dynamic nuclear resonance. J. Chem. Soc. D. 1971, 656–657 (1971).

    Article  Google Scholar 

  30. Download link: https://home.cc.umanitoba.ca/~wolowiec/spinworks/index.html

  31. TopSpin download link: https://www.bruker.com/en/products-and-solutions/mr/nmr-software/topspin.html (2023).

  32. Loening, N. M. & Keeler, J. Temperature accuracy and temperature gradients in solution-state NMR spectrometers. J. Magn. Reson. 159, 55–61 (2002).

    Article  CAS  PubMed  Google Scholar 

  33. Van Geet, A. L. Calibration of methanol nuclear magnetic resonance thermometer at low temperature. Anal. Chem. 42, 679–680 (1970).

    Article  Google Scholar 

  34. Bruker User Manual. (Section 1.3: Temperature Calibration). Available at https://www.bruker.com/protected/en/services/user-manuals/nmr/technical-manuals.html (2023).

Download references

Acknowledgements

We gratefully acknowledge the EPSRC (EP/S024107/1, EP/R005826/1, EP/L015838/1), ERC (DOGMATRON AdG 883786) and Royal Society (RGS\R1\221162) for financial support.

Author information

Authors and Affiliations

  1. School of Chemistry, University of Bristol, Bristol, UK

    Jean-Paul Heeb & Jonathan Clayden

  2. Chemistry Research Laboratory, University of Oxford, Oxford, UK

    Martin D. Smith

  3. School of Natural and Environmental Sciences (Chemistry), Newcastle University, Newcastle Upon Tyne, UK

    Roly J. Armstrong

Authors
  1. Jean-Paul Heeb
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jonathan Clayden
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Martin D. Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Roly J. Armstrong
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

The manuscript was jointly conceived and written through the contributions of all authors.

Corresponding authors

Correspondence to Jonathan Clayden, Martin D. Smith or Roly J. Armstrong.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature Protocols thanks Zhenhua Gu, Osamu Kitagawa and Bingfeng Shi for their contribution to the peer review of this work.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Related links

Key references using this protocol

Costil, R. et al. J. Angew. Chem. Int. Ed. Engl. 59, 18670–18678 (2020): https://doi.org/10.1002/anie.202007595

Staniland, S. et al. Angew. Chem. Int. Ed. Engl. 55, 10755–10759 (2016): https://doi.org/10.1002/anie.201605486

Jolliffe, J. D. et al. Nat. Chem. 9, 558–562 (2017): https://doi.org/10.1038/nchem.2710

Armstrong, R. J. & Smith, M. D. Angew. Chem. Int. Ed. Engl. 53, 12822–12826 (2014): https://doi.org/10.1002/anie.201408205

Surgenor, R. R. et al. Nat. Chem. 15, 357–365 (2023): https://doi.org/10.1038/s41557-022-01095-9

Supplementary information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Heeb, JP., Clayden, J., Smith, M.D. et al. Interrogating the configurational stability of atropisomers. Nat Protoc 18, 2745–2771 (2023). https://doi.org/10.1038/s41596-023-00859-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41596-023-00859-y

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing