Figure 5.

HPLC analysis of click release reaction between Ac₄ManNTz and TCO-Dox. (A) Mechanism of the iEDDA reaction mediated release. Ac₄ManNTz react with TCO-Dox through [4 + 2] cycloaddition to form dihydropyridazine which subsequently undergoes rearrangement and elimination that ultimately liberate the Dox. (B) Representative curves from high-pressure liquid chromatography (HPLC) analysis show cumulative release of Dox after mixing Ac₄ManNTz (40 μmol/L) with TCO-Dox (20 μmol/L) in PBS buffer for 30 min, 1, 2, 4 h. (C) The quantification of released Dox as a function of time. Data are averages ± SEM, n = 3.