Alexander Schiller

Molecular Logic with a Saccharide Probe on the Few-Molecules Level

In this Communication we describe a two-component saccharide probe with logic capability. The combination of a boronic acid-appended viologen and perylene diimide was able to perform a complementary implication/not implication logic function. Fluorescence quenching and recovery with fructose was analyzed with fluorescence correlation spectroscopy on the level of a few molecules of the reporting dye.

Light-triggered CO release from nanoporous non-wovens

The water insoluble and photoactive CO releasing molecule dimanganese decacarbonyl (CORM-1) has been non-covalently embedded into poly(l-lactide-co-d/l-lactide) fibers via electrospinning to enable bioavailability and water accessibility of CORM-1. SEM images of the resulting hybrid non-wovens reveal a nanoporous fiber morphology. Slight CO release from the CORM-1 in the electrospinning process induces nanoporosity. IR spectra show the same set of carbonyl bands for the CORM-1 precursor and the non-woven. When the material was exposed to light (365-480 nm), CO release from the incorporated CORM-1 was measured via heterogeneous myoglobin assay, a portable CO electrode and an IR gas cuvette. The CO release rate was wavelength dependent. Irradiation at 365 nm resulted in four times faster release than at 480 nm. 3.4 μmol of CO per mg non-woven can be generated. Mouse fibroblast 3T3 cells were used to show that the hybrid material is non-toxic in the darkness and strongly photocytotoxic when light is applied.

Multiwell plates loaded with fluorescent hydrogel sensors for measuring pH and glucose concentration

Fluorescent hydrogels were polymerized directly in multi-well plates at ambient temperature and in the presence of air, producing sensors for measuring pH and glucose concentration. The plates were rapidly analyzed using a fluorescence plate reader. Multiwell pH sensors with good reproducibility among different wells and a dynamic range from pH 6 to 9 were prepared by incorporating a polymerizable pH sensitive fluorophore in the hydrogel. Non-enzymatic glucose sensors comprising a boronic acid-appended fluorescence quencher together with an aminopyrene fluorophore were prepared in a matter of hours in multiwell plates. The sensors showed good reproducibility in response to solutions of glucose at physiological pH. Dried glucose sensors rehydrated with analyte solution performed similarly to freshly prepared hydrogels. The loaded plates are designed for use in high throughput screening applications. Plates were prepared using the redox initiator system metabisulfite/persulfate/iron(II) to generate hydrogels of N,N-dimethylacrylamide crosslinked with N,N′-methylenebisacrylamidein situ.

Sugar-based molecular computing by material implication.

A method to integrate an (in principle) unlimited number of molecular logic gates to construct complex circuits is presented. Logic circuits, such as half- or full-adders, can be reinterpreted by using the functional completeness of the implication function (IMP) and the trivial FALSE operation. The molecular gate IMP is represented by a fluorescent boronic acid sugar probe. An external wiring algorithm translates the fluorescent output from one gate into a chemical input for the next gate on microtiter plates. This process is demonstrated on a four-bit full adder.

Enzyme assays with boronic acid appended bipyridinium salts.

In-vitro fluorescent enzyme assays have been developed for sucrose phosphorylase (SPO) and phosphoglucomutase (PGM). These assays make use of a selective carbohydrate sensing system that detects the unlabeled enzymatic products fructose and glucose-6-phosphate. The system comprises 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt as the reporter unit and boronic acid appended viologens as selective receptors with working ranges from 70 microM to 1.0 mM for fructose (SPO) and 190 microM to 2.0 mM for glucose-6-phosphate (PGM). The change in fluorescence can be converted into product concentration, allowing initial reaction velocities and Michaelis-Menten kinetics to be calculated. The assays are also carried out in multiwell plate formats, making them suitable for high-throughput screening of enzyme inhibitors. Rapid PGM inhibition screening is demonstrated with EDTA and LiCl. The PGM assay can also be used for enzyme quantification with a detection limit of 50 ng mL(-1).

Allosteric Indicator Displacement Enzyme Assay for a Cyanogenic Glycoside

Indicator displacement assays (IDAs) represent an elegant approach in supramolecular analytical chemistry. Herein, we report a chemical biosensor for the selective detection of the cyanogenic glycoside amygdalin in aqueous solution. The hybrid sensor consists of the enzyme β-glucosidase and a boronic acid appended viologen together with a fluorescent reporter dye. β-Glucosidase degrades the cyanogenic glycoside amygdalin into hydrogen cyanide, glucose, and benzaldehyde. Only the released cyanide binds at the allosteric site of the receptor (boronic acid) thereby inducing changes in the affinity of a formerly bound fluorescent indicator dye at the other side of the receptor. Thus, the sensing probe performs as allosteric indicator displacement assay (AIDA) for cyanide in water. Interference studies with inorganic anions and glucose revealed that cyanide is solely responsible for the change in the fluorescent signal. DFT calculations on a model compound revealed a 1:1 binding ratio of the boronic acid and cyanide ion. The fluorescent enzyme assay for β-glucosidase uses amygdalin as natural substrate and allows measuring Michaelis-Menten kinetics in microtiter plates. The allosteric indicator displacement assay (AIDA) probe can also be used to detect cyanide traces in commercial amygdalin samples.

Light-triggered NO release from a nanofibrous non-woven

Light-controlled NO delivery systems promise new applications in phototherapies. For this purpose, a ruthenium nitrosyl complex [(2)Ru(NO)(Cl)] with the novel ligand N,N′-(1,2-phenylene)bis(1-methyl-1H-imidazole-2-carboxamide) (2) has been synthesised and characterised in detail. The photoactive {Ru–NO}6 nitrosyl released NO in DMSO upon exposure to low-intensity UV-A light (λ = 366 nm). In order to create an applicable system the water-insoluble [(2)Ru(NO)(Cl)] was embedded into poly(L-lactide-co-D/L-lactide) nanofibrous non-wovens by electrospinning. Exposure of a 25 wt% non-woven of [(2)Ru(NO)(Cl)] to UV-A light resulted in a continuous release of NO into an aqueous solution. The cytoxicity of the non-woven against 3T3 mouse fibroblasts was very low.

Fluorinated Boronic Acid-Appended Pyridinium Salts and 19F NMR Spectroscopy for Diol Sensing

The identification and discrimination of diols is of fundamental importance in medical diagnostics, such as measuring the contents of glucose in the urine of diabetes patients. Diol sensors are often based on fluorophore-appended boronic acids, but these severely lack discriminatory power and their response is one-dimensional. As an alternative strategy, we present the use of fluorinated boronic acid-appended pyridinium salts in combination with 19F NMR spectroscopy. A pool of 59 (bio)analytes was screened, containing monosaccharides, phosphorylated and N-acetylated sugars, polyols, carboxylic acids, nucleotides, and amines. The majority of analytes could be clearly detected and discriminated. In addition, glucose and fructose could be distinguished up to 1:9 molar ratio in mixtures. Crucially, the receptors feature high sensitivity and selectivity and are water-soluble, and their 19F-NMR analyte fingerprint is pH-robust, thereby making them particularly well-suited for medical application. Finally, to demonstrate this applicability, glucose could be detected in synthetic urine samples down to 1 mM using merely a 188 MHz NMR spectrometer.

Sensitization of NO‐Releasing Ruthenium Complexes to Visible Light

We report a combined spectroscopical-theoretical investigation on the photosensitization of transition metal nitrosyl complexes. For this purpose, ruthenium nitrosyl complexes based on tetradentate biscarboxamide ligands were synthesized. A crystal structure analysis of a lithium-based ligand intermediate is described. The Ru complexes have been characterized regarding their photophysical and nitric oxide (NO) releasing properties. Quantum chemical calculations have been performed to unravel the influence of the biscarboxamide ligand frame with respect to the molecular electronic properties of the NO-releasing pathway. A quantitative measure for the ligand design within photosensitized Ru complexes is introduced and evaluated spectroscopically and theoretically by using time-dependent density functional theory.

Going Beyond Continuous Glucose Monitoring with Boronic Acid-Appended Bipyridinium Salts

A two-component sensing system comprising a fluorescent dye and a boronic acid-containing molecule that acts dually as a fluorescence quencher and a saccharide receptor was developed for continuous glucose monitoring in blood. Boronic acid-based bipyridinium salts as tunable receptors have been introduced to increase glucose selectivity over that of other boronate-forming analytes. Powerful solution-phase sensor arrays for neutral and anionic carbohydrates were created with these probes. They can be also used in new label-free fluorescent assays for carbohydrate-modifying enzymes.