This simplified protection method was also successful in producing peptide 1b with reactive DBCO (Figure ). However, while this method successfully generated hydrazide-containing 2a and 1c, side reactions occurred when attempting to react crude 2a or 1c with 6-azidohexanoic acid (Figures ). Copper test strips indicated that Cu was only partially removed during ether precipitation and was likely causing the undesired side reactions during SPAAC. Gratifyingly, HPLC purification appeared to remove the remaining Cu, as pure 1c did not contain Cu (copper test strips), and the peptide successfully coupled with 6-azidohexanoic acid to afford the triazole with minimal side products (Figures ). Inductively coupled plasma mass spectrometry (ICP-MS) confirmed removal of Cu, as several HPLC-purified DBCO peptides contained the same background Cu levels as peptides that were never exposed to the Cu(I) salt (Table ).