A long-lived, highly luminescent Re(I) metal-ligand complex as a biomolecular probe

Abstract

A highly luminescent rhenium (I) metal-ligand complex [Re(bcp)(CO)3(4-COOHPy)](ClO4), where bcp is 2,9-dimethyl-4,7-diphenyl-1,10-phenanthroline and 4-COOHPy is isonicotinic acid, has been synthesized and characterized. High quantum yields (> 0.5) and long excited-state lifetimes (0.3-10 micronseconds) in fluid solutions at room temperature were found for this complex, with remarkable emission sensitivity to microenvironment. This compound also displays highly polarized emission with a maximum anisotropy near 0.3 in the absence of rotational diffusion. This Re complex was conjugated to several biomolecules, including the proteins human serum albumin and bovine immunoglobulin G, as well as an amine-containing lipid. When bound to a protein or lipid, the decay time is near 3 microseconds and the quantum yield is approximately 0.12 in aqueous oxygenated solution at room temperature. This compound's unique spectral properties along with its conjugatability allowed us to utilize it as biomolecular probe in a variety of environments.

FIG. 1.

Molecular structure of [Re(bcp)(CO)₃(4-COOHPy)]⁺.

FIG. 2.

Absorption spectra of [Re(bcp)(CO)₃(4-COOHPy)]⁺, 2,9-dimethyl-4,7-diphenyl-1,10-phenanthroline (bcp), and isonicotinic acid (4-COOHPy) in methanol.

FIG. 3.

Intensity-normalized excitation and emission spectra of [Re(bcp)(CO)₃(4-COOHPy)]⁺ in CHCl₃, CH₃CN, and CH₃OH and in buffer at room temperature. Excitation wavelength was 400 nm. The solid line shows the excitation anisotropy spectrum in 100% glycerol at −60°C, with the emission wavelength tuned to 550 nm. The band pass was 8 nm for all measurements.

FIG. 4.

Temperature-dependent emission spectra (top) and intensity-normalized emission spectra (bottom) of [Re(bcp)(CO)₃⁻(4-COOHPy)]⁺ in 100% glycerol. Excitation was 360 nm with a bandpass of 8 nm.

FIG. 5.

Oxygen-dependent emission spectra of [Re(bcp)(CO)₃⁻(4-COOHPy)]⁺ in CH₃CN at room temperature. Excitation was 400 nm with a bandpass of 8 nm. Inset shows the spectra obtained in air and O₂.

FIG. 6.

Frequency-domain intensity decays of [Re(bcp)(CO)₃⁻(4-COOHPy)]⁺ in methanol. Excitation was 390 nm and a 500-nm cutoff Filter was used to isolate the emission.

FIG. 7.

Oxygen-dependent emission spectra of Re-PE embedded in DPPG vesicles with a mole ratio of Re-PE:DPPG 1:80. Excitation was 400 nm with a bandpass of 8 nm, measured at 20°C.

FIG. 8.

Temperature-dependent emission anisotropy of [Re(bcp)-(CO)₃(4-COOHPy)]⁺ in solutions composed of different ratios of glycerol/H₂O (v/v). Emission was monitored at 550 nm with an excitation wavelength of 400 nm and a bandpass of 8 nm.

FIG. 9.

Excitation spectra and R(l) values for [Re(bcp)(CO)₃⁻ (4-COOHPy)]⁺ in CH₃CN. The R(λ) values were calculated from Eq. [4].

FIG. 10.

Emission anisotropy spectrum of [Re(bcp)(CO)₃⁻(4-COOHPy)]⁺ in 100% glycerol at −06°C. Emission spectrum is shown for comparison. Excitation was 400 nm with a bandpass of 8 nm.

FIG. 11.

Frequency-domain intensity decays of DPPG vesicles la beled with Re-PE at various temperatures. Excitation was 340 nm and a 470-nm cutoff Filter was used to isolate the emission.