Water-soluble 2-hydroxyisophthalamides for sensitization of lanthanide luminescence

Abstract

A series of octadentate ligands featuring the 2-hydroxyisophthalamide (IAM) antenna chromophore to sensitize Tb(III) and Eu(III) luminescence has been prepared and characterized. The length of the alkyl amine scaffold that links the four IAM moieties has been varied to investigate the effect of the ligand backbone on the stability and photophysical properties of the Ln(III) complexes. The amine backbones utilized in this study are N,N,N',N'-tetrakis-(2-aminoethyl)-ethane-1,2-diamine [H(2,2)-], N,N,N',N'-tetrakis-(2-aminoethyl)-propane-1,3-diamine [H(3,2)-], and N,N,N',N'-tetrakis-(2-aminoethyl)-butane-1,4-diamine [H(4,2)-]. These ligands also incorporate methoxyethylene [MOE] groups on each of the IAM chromophores to increase their water solubility. The aqueous ligand protonation constants and Tb(III) and Eu(III) formation constants were determined from solution thermodynamic studies. The resulting values indicate that at physiological pH the Eu(III) and Tb(III) complexes of H(2,2)-IAM-MOE and H(4,2)-IAM-MOE are sufficiently stable to prevent dissociation at nanomolar concentrations. The photophysical measurements for the Tb(III) complexes gave overall quantum yield values of 0.56, 0.39, and 0.52 respectively for the complexes with H(2,2)-IAM-MOE, H(3,2)-IAM-MOE, and H(4,2)-IAM-MOE, while the corresponding Eu(III) complexes displayed significantly weaker luminescence, with quantum yield values of 0.0014, 0.0015, and 0.0058, respectively. Analysis of the steady state Eu(III) emission spectra provides insight into the solution symmetries of the complexes. The combined solubility, stability, and photophysical performance of the Tb(III) complexes in particular make them well suited to serve as the luminescent reporter group in high sensitivity time-resolved fluoroimmunoassays.

Figure 1

Chemical structure of H(2,2)-IAM (top) and monoethylene glycol (MOE)-containing analogs with varied backbones (bottom).

Figure 2

Speciation diagrams for Tb(H(n,2)-IAM-MOE) series (n = 2, 3 and 4 from top to bottom) calculated at 10⁻⁶ M (left column) and 10⁻⁹ M (right column) in aqueous solution.

Figure 3

The observed room temperature absorption (red) and emission spectra (black) of the H(2,2)-IAM-MOE ligand in 0.1 M TRIS buffer at pH 7.4, and corresponding 77 K emission spectra for the Gd(III) complex (blue) in 1:5 (v/v) MeOH:EtOH.

Figure 4

The absorbance (black) and emission spectra (green) (λ_exc = 340 nm) of a 5 µM solution of [Tb(H(2,2)-IAM-MOE)] complex in 0.1 M TRIS buffer at pH 7.4. The corresponding emission spectrum of the [Eu(H(2,2)-IAM-MOE)] complex is shown in red.

Figure 5

Expansion of the emission spectrum for the [Eu(H(2,2)-IAM-MOE)] complex in 0.1 M TRIS buffer (pH 7.4) in the J = 0, 1 region and corresponding fit of the experimental spectrum to four overlapping Gaussians functions.

Scheme 1

H(n,2)-IAM-MOE Ligand Synthesis (n = 2, 3, 4)