# High- and low-field NMR in binary solvent gradients

## TL;DR

This paper introduces a new NMR method using solvent gradients to analyze chemical systems efficiently across different solvent compositions.

## Contribution

A novel NMR technique using solvent composition gradients to study chemical systems in a single experiment.

## Key findings

- The method enables rapid determination of pKa values of APIs in DMSO/water mixtures using high-field NMR.
- Benchtop NMR can transfer resonance assignments between spectra of the same compound in different solvents.
- The approach improves spectral resolution of complex mixtures via differential solubility in solvent gradients.

## Abstract

We introduce composition gradients of the solvent as a powerful new dimension for NMR analysis on both high-field and benchtop instruments. Taking advantage of the differences in the density and miscibility of binary solvent mixtures with different compositions, we layer two solutions at opposite extremes of the compositional range in an NMR tube. The diffusion of the layers into each other establishes a continuous variation in the solvent composition across the sample. Spatially resolved analysis of the sample using either chemical shift imaging (CSI) on high-field NMR instruments or physical movement of the sample (benchtop instruments) enables analysis of chemical systems as a function of the solvent composition. In high field, we determine the pKa of poorly water-soluble active pharmaceutical ingredients (APIs) in a wide range of compositions of dimethylsulfoxide (DMSO)/water and perform accurate extrapolations to aqueous pKa using the Yasuda-Shedlovsky method. We thus condense hours of tedious experiments, where the pKa would be determined separately at each solvent composition, into a single 20 minute experiment. We can also detect the minimum quantity of DMSO required to maintain an API in the freely dissolved state. On a benchtop instrument, we demonstrate how our approach enables the transfer of resonance assignments between spectra of the same compound (asarone) acquired in different solvents (methanol and DMSO). We also show that the method can boost the spectral resolution of complex molecular mixtures (naproxen tablet) via the differential solubility of the components in the two solvents.

Gradients in the solvent composition of NMR samples are generated by layering two solvents of contrasting densities. Spectra are recorded at different compositions by physical movement of the tube (low-field) or chemical shift imaging (high-field).