Application of silver ion in the separation of macrolide antibiotic components by high-speed counter-current chromatography

Three macrolide antibiotic components – ascomycin, tacrolimus and dihydrotacrolimus – were separated and purified by silver ion high-speed counter-current chromatography (HSCCC). The solvent system consisted of n-hexane–tert-butyl methyl ether–methanol–water (1:3:6:5, v/v) and silver nitrate (0.10 mol/l). The silver ion acted as a π-complexing agent with tacrolimus because of its extra side double bond compared with ascomycin and dihydrotacrolimus. This complexation modified the partition coefficient values and the separation factors of the three components. As a result, ascomycin, tacrolimus and dihydrotacrolimus were purified from 150 mg extracted crude sample with purities of 97.6%, 98.7% and 96.5%, respectively, and yields over 80% (including their tautomers). These results cannot be achieved with the same solvent system but without the addition of silver ion.