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Pulsed field gradient nuclear magnetic resonance (PFG-NMR) is a powerful methodology that encodes the microscopic molecular displacements of a substance. Subsequent analysis of the PFG-NMR data then recovers the substance’s molecular self-diffusivity, D, in units of m2 s−1. This chapter is primarily divided into two sections: the first section elucidates the background and theory behind the PFG-NMR experiment and describes mathematically how the molecular self-diffusion coefficient, D, is extracted from PFG-NMR datasets. The latter half of the first section demonstrates how numerical simulations of PFG-NMR data serve as valuable tools to illustrate the effects observed in real PFG-NMR datasets. These effects include multiple-component molecular diffusion, diffusive molecular exchange, and restricted diffusion. The second half of the chapter focuses on the diverse applications of PFG-NMR in late-stage pharmaceutical and biopharmaceutical development and deployment research. It emphasizes specific research aimed at enhancing our scientific comprehension of the principal challenges and questions faced by the pharmaceutical and biopharmaceutical industries.

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