Amido/alkoxy–aryl–aryl–picolinate push–pull antennas for two-photon sensitization of Eu3+ luminescence†

Abstract

Luminescent two-photon (2P) absorbing lanthanide(III) complexes hold great promise for microsocpy imaging of biological samples. Conjugating such a complex to well-chosen cell penetrating peptides (CPP) allows its controlled delivery to the cytosol of live cells. However, alkoxy–phenyl–ethynyl–picolinate, one of the best antennae for 2P sensitization of Eu³⁺, undergoes side reactions at its ethynyl group during peptide synthesis or in biological media and thus cannot be used to create such a conjugate. In this article, we evaluate the effect of substituting the ethynyl group by a phenyl one. We describe the synthesis of conjugates of the TAT CPP with Eu³⁺ complexes featuring amido–phenyl–phenyl–picolinamide, alkoxy–phenyl–phenyl–picolinamide and amido–phenyl–phenyl–picolinate ter-aryl antennae and compare their spectroscopic properties to those of analogues with bi-aryl antennae, including the amido–phenyl–picolinamide already used for 2P live cell imaging. The absorption spectrum of the ter-aryl antennae is red-shifted and better covers the active spectral range for 2P excitation by a Ti-sapphire laser. Among compounds with ter-aryl antennae, those with an amido electron donating group are the most interesting, showing brightness ca. 4 times higher than their bi-aryl counterparts, and similar to the ethynyl-containing antenna. 2P microscopy imaging of live cells incubated with the TAT-Eu³⁺ conjugate and dFFLIPTAT, a non-luminescent CPP that promotes cytosolic delivery, showed diffuse cytosolic staining of the Eu³⁺ probe. The ter-aryl-based probes showed superior performances compared to bi-aryl, with ca. 80% of the cells showing Eu³⁺ staining of the cytosol.