Till Kuehn | Goethe-Universität Frankfurt am Main (original) (raw)

Papers by Till Kuehn

PloS one, 2009

Bacterial meningitis is an acute disease with high mortality that is reduced by early treatment. ... more Bacterial meningitis is an acute disease with high mortality that is reduced by early treatment. Identification of the causative microorganism by culture is sensitive but slow. Large volumes of cerebrospinal fluid (CSF) are required to maximise sensitivity and establish a provisional diagnosis. We have utilised nuclear magnetic resonance (NMR) spectroscopy to rapidly characterise the biochemical profile of CSF from normal rats and animals with pneumococcal or cryptococcal meningitis. Use of a miniaturised capillary NMR system overcame limitations caused by small CSF volumes and low metabolite concentrations. The analysis of the complex NMR spectroscopic data by a supervised statistical classification strategy included major, minor and unidentified metabolites. Reproducible spectral profiles were generated within less than three minutes, and revealed differences in the relative amounts of glucose, lactate, citrate, amino acid residues, acetate and polyols in the three groups. Contrib...

Toxicon, 2007

Bacillus amyloliquefaciens strains isolated from the indoor environment of moisture-damaged build... more Bacillus amyloliquefaciens strains isolated from the indoor environment of moisture-damaged buildings produce a 1197 Da toxin, named amylosin. Nuclear magnetic resonance (NMR) data showed that amylosin contains a chromophoric polyene structure and the amino acids leucine/isoleucine, proline, aspartic acid/asparagine, glutamic acid/glutamine and tyrosine. A quantitation method for amylosin was developed using commercially available amphotericin B as a reference compound and a known concentration of amylosin determined by NMR with the electronic reference to access in vivo concentration (ERETIC) method. Purified amylosin inhibited motility of boar sperm cells at an exposure concentration of 135 nM and hyperpolarized their cell membrane and depolarized their mitochondria at exposure to concentration of 33-67 nM for 10 min. In a 3-d exposure time only 27 nM of amylosin was needed to provoke the same toxicity functions. Amylosin was cytotoxic to feline lung cells at concentrations of <170 nM. Purified amylosin provoked adenosine 5'-triphosphate (ATP)-independent cation influx into isolated rat liver mitochondria (RLM), inducing swelling of the mitochondria at concentrations of 200 nM K(+) or >250 nM Na(+) medium. In the K(+)- or Na(+)-containing medium, amylosin uncoupled RLM, causing oxidation of pyridine nucleotides (PN), loss of the mitochondrial membrane potential, and suppressed ATP synthesis. Purified amylosin produced cation channels in black-lipid membranes (BLMs) with a selectivity K(+)>Na(+) at a concentration of 26 nM, i.e. the same concentration at which amylosin was toxic to boar sperm cells. The amylosin cation channels were cholesterol- and ATP-independent and more effective with K(+) than with Na(+). We propose that the toxicity of amylosin may be due its ionophoric properties, representing the first K(+)/Na(+) channel-forming substance reported from B. amyloliquefaciens.

The Analyst, 2002

The application of a 1 mm TXI (1H/13C/15N) microlitre NMR probe with z-gradient for metabolic pro... more The application of a 1 mm TXI (1H/13C/15N) microlitre NMR probe with z-gradient for metabolic profiling of biofluids is described. The probe was used to provide spectral profiles for rat blood plasma using only approximately 2 microl of fluid with a range of solvent suppression techniques. Using a similar amount of fluid, spectra were obtained from rat and mouse cerebrospinal fluid, demonstrating that the probe could be used to profile rodents metabolically via biofluids previously inaccessible to NMR analysis without the need for termination.

Monatshefte für Chemie - Chemical Monthly, 2011

Abstract The new complete molecular confidence (CMC) concept explores the synergies of the analy... more Abstract The new complete molecular confidence (CMC) concept explores the synergies of the analytical techniques LC–MS and NMR to obtain an estimation of the purity, concentration, and identity of chemical compounds. The high mass accuracy of the MS and MS/MS data provided by the new generation of ESI-TOF and ESI-Q-TOF mass spectrometers provides an accurate determination of molecular weight, which is

Journal of the American Chemical Society, 2000

ABSTRACT The kinetics of protein refolding have been monitored by time-resolved NMR spectroscopy.... more ABSTRACT The kinetics of protein refolding have been monitored by time-resolved NMR spectroscopy. It is shown that refolding of metal binding proteins can be induced by photolysis of photo labile ion chelators, the subsequent release of Ca2+ ions can induce protein folding, and the changes in resonance positions can be monitored by time-resolved NMR spectroscopy. The feasibility of the approach is demonstrated by characterizing the refolding of α-lactalbumin, or protein containing a Ca2+ binding site, unfolded in 4 M urea at pH 7 in the absence of Ca2+. The refolding kinetics of the methyl groups of residues Leu15 and Val21 in the core of the protein have been determined to be mono-exponential with rates of 0.28 s-1 and 0.23 s-1, respectively at 300 K.

Chemistry & Biodiversity, 2005

Introduction. ± Modern spectroscopic methods have largely revolutionized compound identification ... more Introduction. ± Modern spectroscopic methods have largely revolutionized compound identification and tremendously accelerated the pace at which isolated compounds can be identified nowadays. It is hard to believe that 50 years ago compounds still required tedious decomposition into known fragments, or, alternatively, total chemical synthesis to corroborate the proposed structures. However, when compound quantities are very small, total chemical synthesis for independent structure elucidation is still being performed in important cases.

Analytical Chemistry, 2002

A new triple-resonance (TXI) (1H, 13C, 15N) high-resolution nuclear magnetic resonance (NMR) capi... more A new triple-resonance (TXI) (1H, 13C, 15N) high-resolution nuclear magnetic resonance (NMR) capillary probe with 2.5-microL NMR-active sample volume (V(obs)) was built and tested for applications with mass- and volume-limited samples and for coupling of microbore liquid chromatography to NMR. This is the first microliter probe with optimized coil geometry for use with individual capillary tubes with an outer diameter of 1 mm. The 90 degree pulse lengths of the 1-mm microliter probe were below 2 micros for proton, below 8 micros for carbon, and below 20 micros for nitrogen, and a spectral line width at signal half-height below 1 Hz was obtained. Compared to a conventional 5-mm probe, the new 600-MHz 1-mm TXI microliter probe with z-gradient shows an increase in mass sensitivity by a factor of 5, corresponding to a 25-fold reduction in measuring time. The consumption of costly deuterated solvent is reduced by at least 2 orders of magnitude. The 1-mm TXI microliter probe with z-gradient allows the measurement of one-dimensional 1H NMR and two-dimensional heteronuclear NMR spectra with a few nanomoles (micrograms) of compound with high sensitivity, speed, and quality. This is a breakthrough for discrete sample NMR spectroscopy with paramount importance for structure elucidation in natural compound chemistry and metabolic research. It offers also advantages for linking chromatographic methods to NMR in a nindustrial environment. Capillary tube NMR may find new applications in areas where high sample throughput is essential, e.g., in the quality control of large sample arrays from parallel chemistry, screening, and compound depositories. It has the potential to increase the sample throughput by 1 order of magnitude or more if new hardware for fast sample handling and exchange becomes available.

Angewandte Chemie International Edition, 2001

PloS one, 2009

Bacterial meningitis is an acute disease with high mortality that is reduced by early treatment. ... more Bacterial meningitis is an acute disease with high mortality that is reduced by early treatment. Identification of the causative microorganism by culture is sensitive but slow. Large volumes of cerebrospinal fluid (CSF) are required to maximise sensitivity and establish a provisional diagnosis. We have utilised nuclear magnetic resonance (NMR) spectroscopy to rapidly characterise the biochemical profile of CSF from normal rats and animals with pneumococcal or cryptococcal meningitis. Use of a miniaturised capillary NMR system overcame limitations caused by small CSF volumes and low metabolite concentrations. The analysis of the complex NMR spectroscopic data by a supervised statistical classification strategy included major, minor and unidentified metabolites. Reproducible spectral profiles were generated within less than three minutes, and revealed differences in the relative amounts of glucose, lactate, citrate, amino acid residues, acetate and polyols in the three groups. Contrib...

Toxicon, 2007

Bacillus amyloliquefaciens strains isolated from the indoor environment of moisture-damaged build... more Bacillus amyloliquefaciens strains isolated from the indoor environment of moisture-damaged buildings produce a 1197 Da toxin, named amylosin. Nuclear magnetic resonance (NMR) data showed that amylosin contains a chromophoric polyene structure and the amino acids leucine/isoleucine, proline, aspartic acid/asparagine, glutamic acid/glutamine and tyrosine. A quantitation method for amylosin was developed using commercially available amphotericin B as a reference compound and a known concentration of amylosin determined by NMR with the electronic reference to access in vivo concentration (ERETIC) method. Purified amylosin inhibited motility of boar sperm cells at an exposure concentration of 135 nM and hyperpolarized their cell membrane and depolarized their mitochondria at exposure to concentration of 33-67 nM for 10 min. In a 3-d exposure time only 27 nM of amylosin was needed to provoke the same toxicity functions. Amylosin was cytotoxic to feline lung cells at concentrations of <170 nM. Purified amylosin provoked adenosine 5'-triphosphate (ATP)-independent cation influx into isolated rat liver mitochondria (RLM), inducing swelling of the mitochondria at concentrations of 200 nM K(+) or >250 nM Na(+) medium. In the K(+)- or Na(+)-containing medium, amylosin uncoupled RLM, causing oxidation of pyridine nucleotides (PN), loss of the mitochondrial membrane potential, and suppressed ATP synthesis. Purified amylosin produced cation channels in black-lipid membranes (BLMs) with a selectivity K(+)>Na(+) at a concentration of 26 nM, i.e. the same concentration at which amylosin was toxic to boar sperm cells. The amylosin cation channels were cholesterol- and ATP-independent and more effective with K(+) than with Na(+). We propose that the toxicity of amylosin may be due its ionophoric properties, representing the first K(+)/Na(+) channel-forming substance reported from B. amyloliquefaciens.

The Analyst, 2002

The application of a 1 mm TXI (1H/13C/15N) microlitre NMR probe with z-gradient for metabolic pro... more The application of a 1 mm TXI (1H/13C/15N) microlitre NMR probe with z-gradient for metabolic profiling of biofluids is described. The probe was used to provide spectral profiles for rat blood plasma using only approximately 2 microl of fluid with a range of solvent suppression techniques. Using a similar amount of fluid, spectra were obtained from rat and mouse cerebrospinal fluid, demonstrating that the probe could be used to profile rodents metabolically via biofluids previously inaccessible to NMR analysis without the need for termination.

Monatshefte für Chemie - Chemical Monthly, 2011

Abstract The new complete molecular confidence (CMC) concept explores the synergies of the analy... more Abstract The new complete molecular confidence (CMC) concept explores the synergies of the analytical techniques LC–MS and NMR to obtain an estimation of the purity, concentration, and identity of chemical compounds. The high mass accuracy of the MS and MS/MS data provided by the new generation of ESI-TOF and ESI-Q-TOF mass spectrometers provides an accurate determination of molecular weight, which is

Journal of the American Chemical Society, 2000

ABSTRACT The kinetics of protein refolding have been monitored by time-resolved NMR spectroscopy.... more ABSTRACT The kinetics of protein refolding have been monitored by time-resolved NMR spectroscopy. It is shown that refolding of metal binding proteins can be induced by photolysis of photo labile ion chelators, the subsequent release of Ca2+ ions can induce protein folding, and the changes in resonance positions can be monitored by time-resolved NMR spectroscopy. The feasibility of the approach is demonstrated by characterizing the refolding of α-lactalbumin, or protein containing a Ca2+ binding site, unfolded in 4 M urea at pH 7 in the absence of Ca2+. The refolding kinetics of the methyl groups of residues Leu15 and Val21 in the core of the protein have been determined to be mono-exponential with rates of 0.28 s-1 and 0.23 s-1, respectively at 300 K.

Chemistry & Biodiversity, 2005

Introduction. ± Modern spectroscopic methods have largely revolutionized compound identification ... more Introduction. ± Modern spectroscopic methods have largely revolutionized compound identification and tremendously accelerated the pace at which isolated compounds can be identified nowadays. It is hard to believe that 50 years ago compounds still required tedious decomposition into known fragments, or, alternatively, total chemical synthesis to corroborate the proposed structures. However, when compound quantities are very small, total chemical synthesis for independent structure elucidation is still being performed in important cases.

Analytical Chemistry, 2002

A new triple-resonance (TXI) (1H, 13C, 15N) high-resolution nuclear magnetic resonance (NMR) capi... more A new triple-resonance (TXI) (1H, 13C, 15N) high-resolution nuclear magnetic resonance (NMR) capillary probe with 2.5-microL NMR-active sample volume (V(obs)) was built and tested for applications with mass- and volume-limited samples and for coupling of microbore liquid chromatography to NMR. This is the first microliter probe with optimized coil geometry for use with individual capillary tubes with an outer diameter of 1 mm. The 90 degree pulse lengths of the 1-mm microliter probe were below 2 micros for proton, below 8 micros for carbon, and below 20 micros for nitrogen, and a spectral line width at signal half-height below 1 Hz was obtained. Compared to a conventional 5-mm probe, the new 600-MHz 1-mm TXI microliter probe with z-gradient shows an increase in mass sensitivity by a factor of 5, corresponding to a 25-fold reduction in measuring time. The consumption of costly deuterated solvent is reduced by at least 2 orders of magnitude. The 1-mm TXI microliter probe with z-gradient allows the measurement of one-dimensional 1H NMR and two-dimensional heteronuclear NMR spectra with a few nanomoles (micrograms) of compound with high sensitivity, speed, and quality. This is a breakthrough for discrete sample NMR spectroscopy with paramount importance for structure elucidation in natural compound chemistry and metabolic research. It offers also advantages for linking chromatographic methods to NMR in a nindustrial environment. Capillary tube NMR may find new applications in areas where high sample throughput is essential, e.g., in the quality control of large sample arrays from parallel chemistry, screening, and compound depositories. It has the potential to increase the sample throughput by 1 order of magnitude or more if new hardware for fast sample handling and exchange becomes available.

Angewandte Chemie International Edition, 2001