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Selective Suppression of Excipient Signals in 2D 1H-13C Methyl Spectra of Biopharmaceutical Products



Luke W. Arbogast, Frank Delaglio, Joel R. Tolman, John P. Marino


While the use of 1H-13C methyl correlated NMR spectroscopy at natural isotopic abundance has been demonstrated as feasible on protein therapeutics as large as monoclonal antibodies, spectral interference from aliphatic excipients remains a significant obstacle to its widespread application. These signals can cause large baseline artifacts, obscure protein resonances, and cause dynamic range suppression of weak peaks, thus hampering both traditional peak-based spectral analyses as well as emerging chemometric methods of analysis. Here we detail modifications to the 2D 1H-13C gradient-selected HSQC experiment that make use of selective pulsing techniques for targeted removal of interfering excipient signals in spectra of the NISTmAb prepared in several different formulations. This approach is demonstrated to selectively reduce interfering excipient signals while still yielding 2D spectra with only modest loses in protein signal. Furthermore, it is shown that spectral modeling based on the SMILE algorithm can be used to simulate and subtract any residual excipient signals and their attendant artifacts from the resulting 2D NMR spectra.
Journal of Biomolecular Nmr


biotherapeutics, excipients, selective cross-polarization, SMILE, SIERRA


Arbogast, L. , Delaglio, F. , Tolman, J. and Marino, J. (2018), Selective Suppression of Excipient Signals in 2D 1H-13C Methyl Spectra of Biopharmaceutical Products, Journal of Biomolecular Nmr, [online], (Accessed April 14, 2024)
Created November 27, 2018, Updated December 2, 2019