Design and Synthesis of Mg²⁺-Selective Fluoroionophores Based on a Coumarin Derivative and Application for Mg²⁺ Measurement in a Living Cell

Abstract

Novel Mg2+ fluorescent molecular probes (KMG-20-AM and KMG-27-AM; where AM is an acetoxymethyl group) based on a coumarin possessing a charged beta-diketone structure were designed and synthesized. These fluorescent probes produced a red shift from 425 to 445 nm in the absorption spectra after formation of a complex with Mg2+. The fluorescence spectra of these probes also showed a red shift from 485 to 495 nm and an increasing fluorescence intensity after formation of a complex with Mg2+. The optimum experimental conditions were excitation wavelength of 445 nm and a monitored wavelength of 500 nm, where these probes functioned as an indicator showing an image of increasing fluorescence in the presence of Mg2+. These probes showed a "seesaw-type" fluorescent spectral change with the isosbestic point at 480 nm due to the light excitation at 445 nm, which indicates that ratiometry can be used for the measurement. The molecular probes formed a 1:1 complex with Mg2+ and the dissociation constant (Kd) was 10.0 mM for KMG-20. The association constants of the probes with Mg2- were approximately 3 times higher than that with Ca2+, which showed that the selectivity of Mg2+ versus Ca2+ for these probes was over 200 times higher than that for commercially available Mg2+ fluorescent molecular probes such as mag-fura-2, Magnesium Green. As an application of these probes, intracellular fluorescent imaging of Mg2+ was demonstrated using a fluorescent microscope. After the addition of KMG-20-AM and KMG-27-AM into PC12 cells, a strong fluorescence was observed in the cytoplasm and a weak fluorescence in the nuclei region. After treatment with a high-K+ medium, the fluorescence intensity increased due to increasing intracellular Mg2+. The real image of Mg2+ release from the magnesium store was successfully observed with these Mg2+ fluorescent probes.

Dictyostelium Myosin-5b Is a Conditional Processive Motor

“…The use of a new generation of Mg 2ϩ -sensitive fluorescent dyes allows the direct observation of these changes with greater special and temporal resolution. Here, we have shown that similar changes as observed for HL60, HC11, and PC12 cells (49,50) do occur in Dictyostelium (see Fig. 5B and supplemental Movie 2).…”

Coumarin-Derived Fluorescent Chemosensors

“…Two coumarin salen-based sensors 51 and 52 exhibit a pronounced fluorescence enhancement response toward Mg 2+ in the presence of Na + as a synergic trigger (Dong et al, 2011). Coumarin-derived fluorescent molecular probes 53 and 54 can be used for highly selective detection of Mg 2+ versus Ca 2+ by means of monitoring the absorption and fluorescence spectral change (Suzuki et al, 2002). 6 shows the structures of coumarin-derived fluorescent chemosensors for Ag(I).…”

Potential Aluminium(III)- and Gallium(III)-selective Optical Sensors Based on Porphyrazines

“…47 The continuous variation (Job's plot) method was used to determine the binding stoichiometry. [48][49][50] As the maxima of the plots are broadened, pz 1 complexes with Al 3+ and Ga 3+ represent moderate stability ( Fig.…”