Live-cell imaging allows the in vivo analysis of subcellular localisation dynamics of physiological processes with high spatial–temporal resolution. However, only few fluorescent dyes have been custom-designed to facilitate species-specific live-cell imaging approaches in filamentous fungi to date. Therefore, we developed fluorescent dye conjugates based on the sophisticated iron acquisition system of Aspergillus fumigatus by chemical modification of the siderophore triacetylfusarinine C (TAFC). Various fluorophores (FITC, NBD, Ocean Blue, BODIPY 630/650, SiR, TAMRA and Cy5) were conjugated to diacetylfusarinine C (DAFC). Gallium-68 labelling enabled in vitro and in vivo characterisations. LogD, uptake assays and growth assays were performed and complemented by live-cell imaging in different Aspergillus species. Siderophore conjugates were specifically recognised by the TAFC transporter MirB and utilized as an iron source in growth assays. Fluorescence microscopy revealed uptake dynamics and differential subcellular accumulation patterns of all compounds inside fungal hyphae.[Fe]DAFC-NBD and -Ocean Blue accumulated in vacuoles, whereas [Fe]DAFC-BODIPY, -SiR and -Cy5 localised to mitochondria. [Fe]DAFC -FITC showed a uniform cytoplasmic distribution, whereas [Fe]DAFC-TAMRA was not internalised at all. Co-staining experiments with commercially available fluorescent dyes confirmed these findings. Overall, we developed a new class of fluorescent dyes that vary in intracellular fungal targeting , thereby providing novel tools for live-cell imaging applications for Aspergillus fumigatus.

Boron-based dyes are attractive synthetic targets due to their large variability of absorption and emission wavelengths. Through Pictet-Spengler cyclizations, followed by oxidation, π-extended boroquinols have been synthesized. During optimization of the reaction conditions, an unusual dearylation has been found and mechanistically investigated. For two of the synthesized boroquinols, mechanochromic effects with bathochromic shifts up to 50 nm were found upon grinding.

The first technetium-99m tricarbonyl core labelled fluorescent Si-rhodamine and its rhenium analogue for bimodal SPECT- and near-infrared fluorescence imaging is presented.

Background: Silicon rhodamines are of particular interest because of their advantageous dye properties (fluorescence- and biostability, quantum efficiency, tolerance to photobleaching). Therefore, silicon rhodamines find frequent application in STED (stimulated emission depletion) microscopy, as sensor molecules for, e.g., ions and as fluorophores for the optical imaging of tumors. Different strategies were already employed for their synthesis. Because of just three known literature examples in which Suzuki–Miyaura cross couplings gave access to silicon rhodamines in poor to moderate yields, we wanted to improve these first valuable experimental results. Results: The preparation of the xanthene triflate was enhanced and several boron sources were screened to find the optimal coupling partner. After optimization of the palladium catalyst, different substituted boroxines were assessed to explore the scope of the Pd-catalyzed cross-coupling reaction. Conclusions: A number of silicon rhodamines were synthesized under the optimized conditions in up to 91% yield without the necessity of HPLC purification. Moreover, silicon rhodamines functionalized with free acid moieties are directly accessible in contrast to previously described methods.

Radiolabeled fluorescent dyes are decisive for bimodal imaging as well as highly in demand for nuclear- and optical imaging. Silicon-rhodamines (SiRs) show unique near-infrared (NIR) optical properties, large quantum yields and extinction coefficients as well as high photostability. Here, we describe the synthesis, characterization and radiolabeling of novel NIR absorbing and emitting fluorophores from the silicon-rhodamine family for use in optical imaging (OI) combined with positron emission tomography (PET) or single photon emission computed tomography (SPECT), respectively. The presented photostable SiRs were characterized using NMR-, UV-Vis-NIR-spectroscopy and mass spectrometry. Moreover, the radiolabeling conditions using fluorine-18 or iodine-123 were extensively explored. After optimization, the radiofluorinated NIR imaging agents were obtained with radiochemical conversions (RCC) up to 70% and isolated radiochemical yields (RCY) up to 54% at molar activities of g.t. 70 GBq/µmol. Radioiodination delivered RCCs over 92% and allowed to isolate the 123I-labeled product in RCY of 54% at a molar activity of g.t. 7.6 TBq/µmol. The radiofluorinated SiRs exhibit in vitro stabilities g.t. 70% after two hours in human serum. The first described radiolabeled SiRs are a promising step toward their further development as multimodal PET/SPECT-NIR imaging agents for planning and subsequent imaging-guided oncological surgery.

Background: Since their first synthesis, silicon xanthenes and the subsequently developed silicon rhodamines (SiR) gained a lot of attention as attractive fluorescence dyes offering a broad field of application. We aimed for the synthesis of a fluorinable pyridinyl silicon rhodamine for the use in multimodal (PET/OI) medical imaging of mitochondria in cancerous cells. Results: A dihalogenated fluorinatable pyridinyl rhodamine could be successfully synthesized with the high yield of 85% by application of a halogen dance (HD) rearrangement. The near-infrared dye shows a quantum yield of 0.34, comparable to other organelle targeting SiR derivatives and absorbs at 665 nm (εmax = 34 000 M−1cm−1) and emits at 681 nm (τ = 1.9 ns). Using colocalization experiments with MitoTracker® Green FM, we could prove the intrinsic targeting ability to mitochondria in two human cell lines (Pearson coefficient >0.8). The dye is suitable for live cell STED nanoscopy imaging and shows a nontoxic profile which makes it an appropriate candidate for medical imaging. Conclusions: We present a biocompatible, nontoxic, small molecule near-infrared dye with the option of subsequent radiolabelling and excellent optical properties for medical and bioimaging. As a compound with intrinsic mitochondria targeting ability, the radiolabelled analogue can be applied in multimodal (PET/OI) imaging of mitochondria for diagnostic and therapeutic use in, e.g., cancer patients.

Tracers for bimodal optical imaging and positron emission tomography unite multiple advantages in a single molecule. Their tumor-specific uptake can be visualized after their PET activation by radiofluorination via PET/CT or PET/MRI allowing for staging or therapy planning, while their non-radioactive moiety additionally facilitates the visualization of malignant tissue during intraoperative fluorescence-guided surgery or in histological assessments. The silicon-bridged xanthene core offers the opportunity for radiofluorination with SiFA isotope exchange to obtain a small-molecule, PET-activatable NIR dye that can be linked to different target vectors. Herein, we demonstrate for the first time the PET-activation of a fluorinated silicon pyronine, belonging to a class of low-molecular-weight fluorescence dyes with a large Stokes shift (up to 129 nm) and solvent-dependent NIR dye properties, with a successful radiochemical conversion of 70%. The non-fluorinated pyronine precursor is easily accessible by a three-step sequence from commercially starting material with a 12% overall yield. Moreover, a library of seven unusually functionalized (by approximately 15 nm), red-shifted silicon rhodamines were synthesized in three- to four-step sequences and the optical properties of the novel dyes were characterized. It was also shown that the synthesized silicon rhodamine dyes can be easily conjugated by amide bond formation or ‘click-reaction’ approaches.

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