HPLC properties of a new type of nanodiamond (ND) column, based on nonporous glassy carbon core particles coated with polyamine and nanodiamonds in a layer-by-layer (LBL) architecture, were systematically studied and characterised to better understand retention mechanisms¹. Comparisons were made between two batches of these columns denoted as Type I with a thick shell comprising high polyamine/nanodiamond layer count and Type II with a reduced LBL count and  polyamine content used for coating.

Firstly, retention profiles were obtained for a set of model anionic, cationic and ampholytic aromatic analytes as a function of pH revealing substantial anion-exchange properties for both phases attributed to the polyamine protonation, where Type I phase displayed higher overall retention as a consequence of the increased content of polyamine in its shell structure.  The extent of mixed-mode interactions (ion-exchange, hydrophobic) was measured using a broad spectrum of dansylated amino acids (DNS-AA) at varied buffer concentrations (ammonium formate 5-50mM) and organic solvent contents (50%, 70%) with results clearly displaying presence of ion-exchange for the two column types with slopes of logk vs logC approaching 1. Variation in organic modified displayed typical reversed-phase behaviour for more hydrophobic DNS-AAs, meanwhile the order of retention for hydrophilic counterparts alluded to HILIC type retention at play. Ion-exchange capacities were obtained using breakthrough curves revealing capacities of 8 µeq for the Type II and 11 µeq for the Type I columns, this being directly related to higher LBL count of the latter.  Hydrophobic evaluation of the columns produced logP values of 0.40 and 0.39 for the both adsorbents  suggesting low to medium hydrophobicity. Reduction in layers and the use of shorter polyamine polymer chains in the Type II phase increased the overall hydrophobicity and decreased the ion-exchange capacity during the manufacturing process. These ND columns were later subjected to extreme conditions of pH 2 producing a stable, reproducible and baseline separation of 6 DNS-AAs namely Gln, Thr, Ala, Val, Ile and Phe.