Chiral Separation of Biosourced Preservatives: Greenness Evaluation of Liquid and Supercritical Fluid Chromatography...

This study investigated the synthesis and chiral separation of a series of bio-based amide derivatives. Moreover, the greenness of both separation techniques, that is, high-performance liquid chromatography (HPLC) and supercritical fluid chromatography (SFC), was compared. The synthesis employed a two-step, solvent-free, one-pot procedure at 60°C, yielding esters quantitatively and amides with yields of 31%-70%. Chiral separations were performed on polysaccharide stationary phases under the most optimal green conditions. Using a 70/30 heptane/ethanol mobile phase in HPLC, racemate 13 exhibited the highest resolution (R_s = 11.8) and retention time (72.5 min); however, using a 40% ethanol phase reduced both resolution and retention times. SFC, using a 50/50 CO₂/ethanol mobile phase, provided baseline resolution (R_s = 1.51-9.13) for all analytes and significantly reduced analysis times (maximum retention time of 12.6 min for the second eluting enantiomer of racemate 13). Greenness evaluation was carried out using the AGREE and Analytical Method Greenness Score (AMGS) tools. The AGREE score averaged at 0.55 for HPLC and 0.64 for SFC, indicating a slight preference for SFC due to reduced solvent hazards and waste. The AMGS scores confirmed this trend: HPLC showed an average of 40.9 (up to 128 for racemate 13), whereas SFC averaged 14.9 (maximum 31.1). Instrument energy consumption was the dominant contributor to the AMGS score for both techniques, accounting for over 95% of the score for HPLC and over 93% for SFC. For racemate 13, this proportion increased to 99% and 96%, respectively.