Guilty by dissociation: Part B: evaluation of Supercritical Fluid Chromatography (SFC-UV) for the analysis of regioisomeric diphenidine-derived Novel Psychoactive Substances (NPS).

Supercritical fluid chromatography (SFC) with a carbon dioxide and ammonium acetate gradient separates 31 novel diphenidine-derived psychoactive substances, including regioisomers, in under 10 minutes. Different stationary phases (acidic, neutral, basic) produce medium to large selectivity differences between isomers. Acidic silica phases retain diphenidines longer via electrostatic attraction, while basic phases reduce retention via repulsion. Baseline separation is achieved for six of eight substituted groups on a simple silica column. As halo-substituent size increases, resolution between ortho- and meta-isomers decreases, causing co-elution of ortho- and meta-bromodiphenidines. Elution orders differ from reversed-phase UHPLC, providing orthogonal separation, with hydrophilic compounds better retained on SFC columns.

Study at a glance

Abstract

Supercritical Fluid Chromatography (SFC-UV) employing a carbon dioxide (CO2) and 10 mM ammonium acetate in MeOH-water (95:5 v/v) gradient provides a rapid analysis (tG <10 min) of 31 novel, regioisomeric diphenidine-derived psychoactive substances, on a range of stationary phases of differing polarity. Medium to large selectivity differences between regioisomers, were observed on the acidic, neutral and basic SFC phases. For individual substituted ortho-, meta- and para-isomers, the same elution order was observed irrespective of the nature of the stationary phase. The acidic silica stationary phases yielded longer retention of the diphenidines via electrostatic attraction, whereas the basic phases resulted in shorter retention via electrostatic repulsion. SFC effected baseline separation of seven of the eight substituted groups of ortho-, meta- and para-diphenidines evaluated on a range of stationary phases. A simple silica phase achieved baseline separation of six of the regioisomeric substituted diphenidines. As the size of the halo-substituent increased, the resolution between ortho-/meta-isomers decreased, resulting in co-elution of the ortho- and meta-bromodiphenidines. Fluphenidines and chlorodiphenidines generated an elution order of meta- < ortho- < para- whereas an elution order switch was observed for the iodophenidines. This contrasted with RP-UHPLC where the elution order for the fluphenidines and iodophenidines was para- < ortho- < meta- and para- < meta- < ortho- respectively. An orthogonal elution order of diphenidines was demonstrated between the RP-UHPLC and SFC stationary phases due to the polarity differences between the separation modes. In general, hydrophilic compounds, which were poorly retained on a C18 reverse phase column, were well retained on SFC columns.