Fluorescent Arylphosphonic Acids: Synergic Interactions between Bone and the Fluorescent Core

Abstract

Herein, we report the third generation of fluorescent probes (arylphosphonic acids) to target calcifications, particularly hydroxyapatite (HAP). In this study, we use highly conjugated porphyrin‐based arylphosphonic acids and their diesters, namely 5,10,15,20‐tetrakis[m‐(diethoxyphosphoryl)phenyl]porphyrin (m‐H8TPPA‐OEt8) and 5,10,15,20‐tetrakis [m‐phenylphosphonic acid]porphyrin (m‐H8TPPA), in comparison with their positional isomers 5,10,15,20‐tetrakis[p‐(diisopropoxyphosphoryl)phenyl]porphyrin (p‐H8TPPA‐iPr8) and 5,10,15,20‐tetrakis [p‐phenylphosphonic acid]porphyrin (p‐H8TPPA), which have phosphonic acid units bonded to sp2 carbon atoms of the fluorescent core. The conjugation of the fluorescent core is thus extended to the (HAP) through sp2‐bonded −PO3H2 units, which generates increased fluorescence upon HAP binding. The resulting fluorescent probes are highly sensitive towards the HAP in rat bone sections. The designed probes are readily taken up by cells. Due to the lower reported toxicity of (p‐H8TPPA), these probes could find applications in monitoring bone resorption or adsorption, or imaging vascular or soft tissue calcifications for breast cancer diagnosis etc.

Arylphosphonate‐Tethered Porphyrins: Fluorescence Silencing Speaks a Metal Language in Living Enterocytes**

“…Ca 2+ or Mg 2+ treatment did not shift the position of the Soret maximum, but slightly raised the intensity of the absorption spectrum causing a higher fluorescence, characteristic for non‐ratiometric metal‐ON fluorescence sensors. This observation confirms our recent results of H 8 TPPA and m ‐H 8 TPPA fluorescence enhancement in the presence of calcium‐hydroxyapatite phases or bones [36] . Mn 2+ , and far more pronounced Pb 2+ , narrowed non‐ratiometrically the photon absorbing properties of p ‐H 8 TPPA and m ‐H 8 TPPA, thus their fluorescence dropped markedly.…”

“…p-H 8 TPPA (Figure 1) and m-H 8 TPPA (Figure S1 in the Supporting information), were pretreated with various bivalent metal ions at a pH close to the cellular milieu (pH 7.4) before recording absorption and fluorescence spectra. The visible absorption of both sensors in the buffer-controlled treatment showed maxima in the range of 380-430 nm, with a typical Soret peak at around 416 nm, already observed in our recent studies [36,44] and also by others. [45] Ca 2 + or Mg 2 + treatment did not shift the position of the Soret maximum, but slightly raised the intensity of the absorption spectrum causing a higher fluorescence, characteristic for nonratiometric metal-ON fluorescence sensors.…”

“…This observation confirms our recent results of H 8 TPPA and m-H 8 TPPA fluorescence enhancement in the presence of calcium-hydroxyapatite phases or bones. [36] Mn 2 + , and far more pronounced Pb 2 + , narrowed non-ratiometrically the photon absorbing properties of p-H 8 TPPA and m-H 8 TPPA, thus their fluorescence dropped markedly.…”

“…In contrast, sp 3 bonded metal sensing units could carry the fluorescent tag to a targeted area without altering the fluorescent signal upon metal binding. We have recently shown that such a design strategy in extending the conjugation of the fluorescent core allows direct electronic interactions with rat bone sections, in return producing turn up fluorescence upon hydroxyapatite binding using p-H 8 TPPA and m-H 8 TPPA fluorescent probes, [36] with toxicity studies indicating that p-H 8 TPPA was well tolerated by an intestinal cell line. [37] Phenylphosphonic acid tethers provide 1.7 and 7.4 pK a 1 and pK a 2 values, respectively, [38,39] and each of the phenylphosphonic acid tethers are expected to provide À 2 negative charge at physiological pH.…”

Vascular Calcification in Rodent Models—Keeping Track with an Extented Method Assortment

“…The application was possible both ex vivo and in vitro. Several other dyes targeting hydroxyapatite have been reported, including, for example, the long-established dye calcein [ 122 , 123 ], fluorescence-labeled osteocalcin [ 124 ] and an arylphosphonic acid-based dye [ 125 ]. The development of new fluorescent dyes may offer several advantages over von Kossa and alizarin red staining.…”