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- Fluorescence (4)
- Fluoreszenz (3)
- Enantioselectivity (2)
- Enantioselektivität (2)
- Molecular imprinting (2)
- Molecularly imprinted polymers (2)
- Aminosäuren (1)
- Biomarker (1)
- Chromatography (1)
- Core-shell nanoparticles (1)
Organisationseinheit der BAM
A 2,3-diaminophenazine bis-urea fluorescent probe monomer (1) was developed. It responds to phenylphosphate and phosphorylated amino acids in a ratiometric fashion with enhanced fluorescence accompanied by the development of a redshifted emission band arising from an excited-state proton transfer (ESPT) process in the hydrogen-bonded probe/analyte complex. The two urea groups of 1 form a cleft-like binding pocket (Kb>10^10 L^2 mol^-2 for 1:2 complex). Imprinting of 1 in presence of ethyl ester- and fluorenylmethyloxycarbonyl (Fmoc)-protected phosphorylated tyrosine (Fmoc-pTyr-OEt) as the template, methacrylamide as co-monomer, and ethyleneglycol dimethacrylate as crosslinker gave few-nanometer-thick molecularly imprinted polymer (MIP) shells on silica core microparticles with excellent selectivity for the template in a buffered biphasic assay. The supramolecular recognition Features were established by spectroscopic and NMR studies. Rational screening of comonomers and cross-linkers allowed to single out the best performing MIP components, giving significant imprinting factors (IF>3.5) while retaining ESPT emission and the ratiometric response in the thin polymer shell. Combination of the bead-based detection scheme with the phase-transfer assay dramatically improved the IF to 15.9, allowing sensitive determination of the analyte directly in aqueous media.
Integration of a polymerisable fluorescent urea-nitrobenzoxadiazole monomer 1 into a few-nanometre thin molecularly imprinted polymer (MIP) shell coated onto a 300 nm silica core with the tetrabutylammonium salt of N-carbobenzyloxy-L-phenylalanine (Cbz- or Z-ʟ-Phe) as the imprinted template yields core/shell MIP nanoparticles that respond to the designated analyte with a strong fluorescence increase in the visible spectral range. The system shows promising imprinting and enantioselectivity factors of 3.6 and 1.9 when benzylmethylacrylate (BMA) is used as co-monomer and ethylene glycol dimethacrylate (EDMA) as cross-linker. A second, naphthalimide-based fluorescent urea 2 is employed in the preparation of MIP thin-films and monolithic polymers for separation techniques. Spectroscopic and chromatographic studies of the MIPs with the analytes Z-ʟ-Phe, Z-ʟ-glutamic acid (Z-ʟ-Glu) and penicilin G (PenG) revealed the (enantio)selective discrimination behaviour of these materials.
A naphthalimide-based fluorescent indicator monomer 1 for the integration into chromo- and fluorogenic molecularly imprinted polymers (MIPs) was synthesized and characterized. The monomer was equipped with a urea binding site to respond to carboxylate-containing guests with absorption and fluorescence changes, namely a bathochromic shift in absorption and fluorescence quenching. Detailed spectroscopic analyses of the title compound and various models revealed the signaling mechanism. Titration studies employing benzoate and Z-ʟ-phenylalanine (Z-ʟ-Phe) suggest that indicator monomers such as the title compound undergo a mixture of deprotonation and complex formation in the presence of benzoate but yield hydrogen-bonded complexes, which are desirable for the molecular imprinting process, with weakly basic guests like Z-ʟ-Phe. Compound 1 could be successfully employed in the synthesis of monolithic and thin-film MIPs against Z-ʟ-Phe, Z-L-glutamic acid, and penicillin G. Chromatographic assessment of the selectivity features of the monoliths revealed enantioselective discrimination and clear imprinting effects. Immobilized on glass coverslips, the thin-film MIPs of 1 displayed a clear signaling behavior with a pronounced enantioselective fluorescence quenching dependence and a promising discrimination against cross-analytes.
The expression of cell surface glycans terminating with sialic acid (SA) residues has been found to correlate with various disease states there among cancer. We here report a novel strategy for specific fluorescence labeling of such motifs. This is based on sialic acid-imprinted core–shell nanoparticles equipped with nitrobenzoxadiazole (NBD) fluorescent reporter groups allowing environmentally sensitive fluorescence detection at convenient excitation and emission wavelengths. Imprinting was achieved exploiting a hybrid approach combining reversible boronate ester formation between p-vinylphenylboronic acid and SA, the introduction of cationic amine functionalities, and the use of an NBD-appended urea-monomer as a binary hydrogen-bond donor targeting the SA carboxylic acid and OH functionalities. The monomers were grafted from 200 nm RAFT-modified silica core particles using ethylene glycol dimethacrylate (EGDMA) as cross-linker resulting in a shell thickness of ca. 10 nm. The particles displayed strong affinity for SA in methanol/water mixtures (K = 6.6 × 105 M-1 in 2% water, 5.9 × 103 M-1 in 98% water, Bmax ≈ 10 µmol g-1), whereas binding of the competitor glucuronic acid (GA) and other monosaccharides was considerably weaker (K (GA) = 1.8 × 103 M-1 in 98% water). In cell imaging experiments, the particles selectively stained different cell lines in correlation with the SA expression level. This was further verified by enzymatic cleavage of SA and by staining using a FITC labeled SA selective lectin.
Sphingosine-1-phosphate (S1P) is a bioactive sphingo-lipid with a broad range of activities coupled to its role in G-protein coupled receptor signalling. Monitoring of both intra and extra cellular levels of this lipid is challenging due to its low abundance and lack of robust affinity assays or sensors. We here report on fluorescent sensory core-shell molecularly imprinted polymer (MIP) particles responsive to near physiologically relevant levels of S1P and the S1P receptor modulator fingolimod phosphate (FP) in spiked human serum samples. Imprinting was achieved using the tetrabutylammonium (TBA) salt of FP or phosphatidic acid (DPPA·Na) as templates in combination with a polymerizable nitrobenzoxadiazole (NBD)-urea monomer with the dual role of capturing the phospho-anion and signalling its presence. The monomers were grafted from ca 300 nm RAFT-modified silica core particles using ethyleneglycol dimethacrylate (EGDMA) as crosslinker resulting in 10–20 nm thick shells displaying selective fluorescence response to the targeted lipids S1P and DPPA in aqueous buffered media. Potential use of the sensory particles for monitoring S1P in serum was demonstrated on spiked serum samples, proving a linear range of 18–60 μM and a detection limit of 5.6 μM, a value in the same range as the plasma concentration of the biomarker.