Evaporation-free Supercritical Fluid Chromatography-Internal Standard Correction-Solvent Suppression-Quantitative...

Accurate quantification of structurally similar impurities in chiral drugs remains a critical challenge in pharmaceutical quality control. Herein, we established supercritical fluid chromatography-internal standard correction-solvent suppression-quantitative nuclear magnetic resonance (SFC-ISC-SS-qNMR) for the rapid and accurate purity assessment of low-purity quinine (QN). Baseline separation of QN from its chiral isomer quinidine was achieved within 10 min using SFC. The collected eluate, without any nitrogen blowing or freeze-drying, was directly mixed with an IS and a trace amount of deuterium oxide for ¹H qNMR analysis, reducing the total sample preparation time to less than 7 min. An excitation sculpting pulse sequence (zgesgp) was employed to effectively suppress the methanol solvent peak, yielding a flat baseline and an excellent signal-to-noise ratio (S/N > 5000:1). Internal standard correction based on SFC peak area ratios before and after purification eliminated losses and errors during sample preparation. Method validation showed that the determined purity of QN (95.40% ± 0.17%) was in agreement with results obtained by two independent direct qNMR strategies (95.61% ± 0.11% and 95.44% ± 0.23%). Compared with previously reported ISC-high-performance liquid chromatography-qNMR and SFC-ISC-qNMR methods, the proposed approach significantly reduced the sample preparation time, completely avoided the nitrogen blowing or freeze-drying steps, and achieved seamless hyphenation of SFC and qNMR. This strategy provides an efficient, green, and generalizable paradigm for the accurate quantification of low-purity organic compounds in chiral drugs and complex matrices.