Residual chemical shift anisotropy

Residual chemical shift anisotropy (RCSA) is another parameter that is being incorporated into the structure elucidation toolbox. As its name suggests, RCSA is similar to residual dipolar coupling (RDC) in that it requires partial alignment of the sample so that differences between the aligned and unaligned states can be measured.

In normal solution NMR each nucleus gives a signal at a characteristic frequency that depends on the electronic shielding around that nucleus. The cloud of electrons that shields each nucleus is rarely spherical, however, so the amount of shielding depends on the orientation of the electron cloud with respect to the magnetic field. In the isotropic state, where all molecules are tumbling randomly and all orientations of the electron cloud are present, the shielding averages out so that a single peak is observed in the spectrum. If the tumbling is restricted in one direction, then the shielding will be altered and the frequencies of the nuclei and the position of their peaks in the NMR spectrum will change as well.

RCSA measurements can be a powerful tool for structure elucidation because, like RDCs, they provide global constraints, rather than the local ones derived from NOES and scalar coupling. Compared to RDCs, RCSAs are more sensitive because they are derived from a singlet peak rather than a coupled doublet. And unlike RDCs, it is easy to obtain RCSAs from quarternary carbons, which may be of special importance for proton poor structures.

To measure RCSAs it is best to use a single sample and switch between isotropic and anisotropic states as this eliminates the possibility of concentration or solvent differences influencing the chemical shifts. Not all alignment media allow this, but of those that do, stretched or compressed gels are the most amenable to small molecule work. Several groups have developed modified NMR tubes that allow gels to be stretched and compressed^1,2,3, and some are available commercially⁴. Generally the devices use a screw-in piston to force a polymeric gel into a reduced volume NMR tube which is then held in a normal 5mm NMR tube. The gel is soaked in a solution of the sample before being forced into the tube. By changing the extent of compression different degrees of alignment can be obtained.

Most groups that have measured RCSAs on small molecules have used the information to select one structure from a group of possibilities^5,6,7. Typically, RCSAs were recorded on molecules with multiple stereo-centers, then the experimental values were compared against theoretical RCSAs of all the possible configurations calculated using density functional theory. Calculating a quality factor for each configuration allowed the one that best explained the experimental data to be identified⁸. Since the sample preparation methods for RCSAs and RDCs are the same, both parameters can be obtained with the same setup. And in some cases, it was found that both parameters were required to unambiguously determine the structure⁵.

References

1. Hellemann E, Gil RR.
New Stretching Method for Aligning Gels: Its Application to the Measurement Residual Chemical Shift Anisotropies (RCSAs) without the Need for Isotropic Shift Correction.
Chemistry. 2018 Mar 12;24(15):3689-3693

2. Liu Y, Prestegard JH.
A device for the measurement of residual chemical shift anisotropy and residual dipolar coupling in soluble and membrane-ssociated proteins.
J Biomol NMR. 2010 Aug;47(4):249-58

3. Kuchel PW, Chapman BE, Müller N, Bubb WA, Philp DJ, Torres AM
Apparatus for rapid adjustment of the degree of alignment of NMR samples in aqueous media: verification with residual quadrupolar splittings in (23)Na and (133)Cs spectra.
J Magn Reson. 2006 Jun;180(2):256-65

4. New Era: Gel NMR Sample Tubes and Accessories

5. Hallwass F, Schmidt M, Sun H, Mazur A, Kummerlöwe G, Luy B, Navarro-Vázquez A, Griesinger C, Reinscheid UM
Residual chemical shift anisotropy (RCSA): a tool for the analysis of the configuration of small molecules.
Angew Chem Int Ed Engl. 2011 Sep 26;50(40):9487-90

6. Nath N, Schmidt M, Gil RR, Williamson RT, Martin GE, Navarro-Vázquez A, Griesinger C, Liu Y
Determination of Relative Configuration from Residual Chemical Shift Anisotropy.
J Am Chem Soc. 2016 Aug 3;138(30):9548-56

7. Liu Y, Saurí J, Mevers E, Peczuh MW, Hiemstra H, Clardy J, Martin GE, Williamson RT
Unequivocal determination of complex molecular structures using anisotropic NMR measurements.
Science. 2017 Apr 7;356(6333). pii: eaam5349.

8. Liu Y, Navarro-Vázquez A, Gil RR, Griesinger C, Martin GE, Williamson RT
Application of anisotropic NMR parameters to the confirmation of molecular structure.
Nat Protoc. 2019 Jan;14(1):217-247