Elution behavior of drugs in high-speed counter-current chromatography using on-column complexation with metal ions

In this study, determination of (nitrogen containing) drugs by on-column complexation with metal ions in high-speed counter-current chromatography (HSCCC) was investigated. Bromazepam (BMP) was strongly retained in the organic upper stationary phase (UP) of the two-phase solvent system composed of tert-butyl methyl ether-acetonitrile–water (2:2:3, v/v/v) by eluting the aqueous lower mobile phase (LP) at a flow rate of 2 mL min⁻¹. On the other hand, BMP (200 µg mL⁻¹) was eluted faster without retention to the organic UP with the two-phase system containing 100 μg mL⁻¹ of copper ions (CuCl₂) because a very polar BMP-Cu²⁺ complex was immediately formed in the aqueous LP. The dramatic change in the retention behavior of BMP resulted from on-column complexation. The on-column complexation in HSCCC was further investigated for five (nitrogen containing) drugs and seven metal ions. In the result, tizanidine and phentolamine formed complexes with Al³⁺, Fe²⁺, Co²⁺, Ni²⁺, Cu²⁺, and Zn²⁺, ambroxol formed complexes with Al³⁺, Fe²⁺, and Cu²⁺, but voriconazole formed no complexes with all metal ions tested.