Fluorescence Sensors for Lithium Ions and Small Peptides

Simple but powerful chemosensors were developed by taking advantage of self-assembly processes. First, we will describe a turn-on fluorescent sensor for the pharmacologically important lithium ion. The sensor can be used in purely aqueous solution, and it displays a very high selectivity for lithium over sodium and magnesium ions. It is based on a metallacrown receptor unit, which can be assembled in situ from simple building blocks. Furthermore, we will describe a cross-reactive sensor array for the detection of small peptides. The individual sensors of the array are formed by mixing metal complexes with fluorescent dyes.

Metal-Based Chemosensors for Important Bioanalytes

Sébastien Rochat

This work describes different strategies for the detection of biologically relevant analytes. The main goal was to achieve sensing in water, which is a prerequisite for application to real biological samples. In several cases, we could demonstrate that our systems were also working in blood serum, a notoriously complex medium containing proteins, sugars and salts, which can potentially interfere with our sensors. Our first analyte of choice was the lithium cation. Owing to its therapeutic uses in the field of psychology, it represents an appealing target, and an efficient chemosensor for lithium ions may find numerous uses in biomedical investigations. The first strategy that will be presented involves the use of organometallic macrocycles carrying fluorescent reporting units. These compounds were found to be potent ionophores and display a very high selectivity for lithium ions. Upon careful choice of constituent building blocks, a chemosensor displaying good affinity, selectivity, and solubility was obtained. The sensor allowed the detection of lithium ions in water and in human serum by simple fluorescence measurements (Chapter 2). Other approaches for the detection of lithium ions will be presented in Chapter 3. Instead of covalently attaching fluorescent dyes to the lithium binding unit, fluorophores that can bind in a non-covalent fashion were employed. Assays based on easily available constituents were devised. They allowed the sensing of lithium ions in the sub-millimolar concentration range. In Chapter 4, the pattern-based sensing of micromolar concentrations of small peptides in water will be described. Small peptides are ubiquitous in biology, and they are involved in many essential processes such as neurotransmission, blood pressure regulation or oxidative stress mitigation. It will be demonstrated that their selective detection can be accomplished by arrays of differential sensors: instead of building a selective sensor for a given analyte, a series of cross-reactive, unsophisticated sensors could be employed. Their differential response (fingerprint) for various analytes was interpreted via statistical methods. As a result, low micromolar concentrations of many short peptides (as well as mixtures thereof) could be measured in water and in human serum samples. In the last part (Chapter 5), the sensing properties of a sensor array will be compared with those of a dynamic combinatorial library (DCL). The DCL contains the same elements as the sensor array, but they are mixed in one pot. It is shown that the DCL can outperform the sensor array in many situations, but that an excessive complexity can be detrimental to its resolution.

EPFL2010

Metal-based chemosensors for amino acids, peptides, and nucleotides

Andrey Buryak

An organometallic 4d transition metal complex [CpRhCl2]2, together with commercially available dyes, was used to construct indicator displacement assays (IDAs) for the detection of peptides, amino acids, and nucleotides. The combination of the CpRh complex with the dye azophloxine was found to form a chemosensing ensemble for the sequence-selective detection of histidine- and methionine-containing peptides in water at neutral pH. A strong interaction of the rhodium complex with peptides bearing histidine or methionine residue in the position 1 or 2 allowed to detect these peptides down to a concentration of 0.3 µM. The same organometallic complex and three commercially available dyes were employed to compose an array of IDA chemosensors for the detection of natural amino acids. We found that the variation of the pH can effectively be used to modulate the selectivity of an IDA sensor. An excellent discrimination of 20 amino acids was achieved. The combination of [Cp*RhCl2]2 with the dyes gallocyanine, evans blue, and mordant yellow 10 forms a multicomponent indicator displacement assay (MIDA), which can be used to sense low millimolar concentrations of nucleotides and the pyrophosphate anion in buffered aqueous solution. Moreover, the MIDA allows to simultaneously determine the concentrations of adenosine triphosphate and pyrophosphate/cyclic adenosine monophosphate with a single UV-Vis measurement. In the second part of our work we investigated whether dynamic combinatorial libraries (DCLs) can be used for sensing purposes. We found that the dyes arsenazo I, methylcalcein blue, and glycine cresol red, in combination with the metal salts CuCl2 and NiCl2, form a DCL of differently colored metal-dye complexes. The addition of an analyte able to interact with a member(s) of such a library, results in a re-equilibraion easily detectable by UV-Vis spectroscopy. Using dipeptides as analytes we demonstrated that this way of sensing is very effective – even closely related peptides such as regio- (Ala-Phe vs. Phe-Ala) and stereo- (Phe-Ala vs. D-Phe-Ala) isomers can easily be discriminated. Additionally, we found that the identity of the best sensor depends on the problem to be addressed.

EPFL2007

Cross-Reactive Sensor Arrays for the Detection of Peptides in Aqueous Solution by Fluorescence Spectroscopy

Jie Gao, Sébastien Rochat, Kay Severin, Friederike Gundula Marie Zaubitzer

A simple but powerful method for the sensing of peptides in aqueous solution has been developed. The transition-metal complexes [PdCl2(en)], [{RhCl2Cp*}2], and [{RuCl2(p-cymene)} 2] were combined with six different fluorescent dyes to build a cross-reactive sensor array. The fluorescence response of the individual sensor units was based on competitive complexation reactions between the peptide analytes and the fluorescent dyes. The collective response of the sensor array in a time-resolved fashion was used as an input for multivariate analyses. A sensor array comprised of only six metal-dye combinations was able to differentiate ten different dipeptides in buffered aqueous solution at a concentration of 50 uM. Furthermore, the cross-reactive sensor could be used to obtain information about the identity and the quantity of the pharmacologically interesting dipeptides carnosine and homocarnosine in a complex biological matrix, such as deproteinized human blood serum. The sensor array was also able to sense longer peptides, which was demonstrated by differentiating mixtures of the nonapeptide bradykinin and the decapeptide kallidin.