Effect of the length of polyoxyethylene substituents on luminescent bimetallic lanthanide bioprobes

The new homoditopic ligand $H_2L^{C2'}$ self-assembles with lanthanide ions ($Ln^{III}$) to yield neutral bimetallic helicates of overall composition $[Ln_2(L^{C2'})_3]$; it is fitted with two hexakis(oxyethylene) chains to test their effects on the thermodynamic, photophysical and biochemical properties of these complexes, with particular emphasis on their uptake by living cells. At physiological pH and under stoichiometric conditions, the conditional stability constants $\log \beta_{23}$ are around 28 resulting in the speciation of the $Eu^{III}$ helicate being > 92% for a total ligand concentration of 1 mM. The ligand triplet state features adequate energy (0- phonon transition at $\approx 21 800 cm^{-1}$) for sensitising the luminescence of $Eu^{III}$ (Q = 19%) and $Tb^{III}$ (Q = 10%) in aerated water at pH 7.4. The $Eu(^5D_0)$ emission spectrum and lifetime (2.43 ms) are characteristic of a species with pseudo-$D_3$ symmetry and without bound water in the inner coordination sphere. The viability of HeLa cancerous cells is unaffected when incubated with up to 500 microM $[Eu_2(L^{C2'})_3]$ during 24 h. The $Eu^{III}$ helicate permeates into the cytoplasm of these cells by endocytosis and remains essentially undissociated, despite a low intracellular concentration of $0.28 \mu M$. In addition, the leakage of the $Eu^{III}$ helicate out of HeLa cells is very minimal over long periods of time. With respect to similar complexes with ligands bearing shorter tris(oxyethylene) chains, no substantial changes are observed, which opens the way for targeting experiments. This study also demonstrates that the $[Ln_2(L^{CX})_3]$ helicates are fairly robust entities since their core is unaffected by the substitution in the pyridine 4-position.