High-throughput Nuclear Magnetic Resonance Research Articles

Metabolic signatures of insulin resistance in 7,098 young adults.

Metabolite associations with insulin resistance were studied in 7,098 young Finns (age 31 ± 3 years; 52% women) to elucidate underlying metabolic pathways. Insulin resistance was assessed by the homeostasis model (HOMA-IR) and circulating metabolites quantified by high-throughput nuclear magnetic resonance spectroscopy in two population-based cohorts. Associations were analyzed using regression models adjusted for age, waist, and standard lipids. Branched-chain and aromatic amino acids, gluconeogenesis intermediates, ketone bodies, and fatty acid composition and saturation were associated with HOMA-IR (P < 0.0005 for 20 metabolite measures). Leu, Ile, Val, and Tyr displayed sex- and obesity-dependent interactions, with associations being significant for women only if they were abdominally obese. Origins of fasting metabolite levels were studied with dietary and physical activity data. Here, protein energy intake was associated with Val, Phe, Tyr, and Gln but not insulin resistance index. We further tested if 12 genetic variants regulating the metabolites also contributed to insulin resistance. The genetic determinants of metabolite levels were not associated with HOMA-IR, with the exception of a variant in GCKR associated with 12 metabolites, including amino acids (P < 0.0005). Nonetheless, metabolic signatures extending beyond obesity and lipid abnormalities reflected the degree of insulin resistance evidenced in young, normoglycemic adults with sex-specific fingerprints.

An inexpensive high-throughput nuclear magnetic resonance tube cleaning apparatus

Large-scale nuclear magnetic resonance (NMR) tube cleaning is currently a bottleneck in high-throughput NMR ligand affinity screens. Expensive alternatives include discarding the NMR tubes after a single use (∼US $2–$8/tube), using commercial NMR tube cleaners (∼$15,000), and abandoning NMR tubes for flow probe technology (∼$75,000). Instead, we describe a relatively inexpensive (∼$400) and easily constructed apparatus that can clean 180 NMR tubes per hour while using a modest amount of solvent. The application of this apparatus significantly shortens the time to recycle NMR tubes while avoiding cross-contamination and tube damage.

Mixture analysis by NMR as applied to fruit juice quality control

Nuclear magnetic resonance (NMR) spectroscopy is rapidly gaining importance in mixture analysis, originally driven by the pharmaceutical and nowadays also by clinical applications within metabonomics. Quality control of food-related material has very similar requirements, as it also deals with mixtures, and many of the compounds found in body fluids are analyzed as well. NMR allows analysis in two ways within one experiment: namely, targeted and untargeted. Targeted stands for the safe identification and consequent quantification of individual compounds, whereas untargeted means the detection of all deviations visible by NMR using statistical analysis based on normality models. Very important is the stability and reproducibility of the NMR instrumentation used, and this means inherent minimized system internal variance. NMR is especially suited for such requirements, as it allows detection of the smallest concentration changes of many metabolites simultaneously. High-throughput flow-injection NMR as the basis for fruit juice screening allows low cost per sample and delivers substantially more relevant information than any other method and is probably the only method to produce such results.

High-throughput serum NMR metabonomics for cost-effective holistic studies on systemic metabolism

A high-throughput proton (1H) nuclear magnetic resonance (NMR) metabonomics approach is introduced to characterise systemic metabolic phenotypes. The methodology combines two molecular windows that contain the majority of the metabolic information available by 1H NMR from native serum, e.g. serum lipids, lipoprotein subclasses as well as various low-molecular-weight metabolites. The experimentation is robotics-controlled and fully automated with a capacity of about 150-180 samples in 24 h. To the best of our knowledge, the presented set-up is unique in the sense of experimental high-throughput, cost-effectiveness, and automated multi-metabolic data analyses. As an example, we demonstrate that the NMR data as such reveal associations between systemic metabolic phenotypes and the metabolic syndrome (n = 4407). The high-throughput of up to 50,000 serum samples per year is also paving the way for this technology in large-scale clinical and epidemiological studies. In contradiction to single 'biomarkers', the application of this holistic NMR approach and the integrated computational methods provides a data-driven systems biology approach to biomedical research.

High-Throughput Nuclear Magnetic Resonance Metabolomic Footprinting for Tissue Engineering

We report a high-throughput (HTP) nuclear magnetic resonance (NMR) method for analysis of media components and a metabolic schematic to help easily interpret the data. Spin-lattice relaxation values and concentrations were measured for 19 components and 2 internal referencing agents in pure and 2-day conditioned, hormonally defined media from a 3-dimensional (3D) multicoaxial human bioartificial liver (BAL). The (1)H NMR spectral signal-to-noise ratio is 21 for 0.16 mM alanine in medium and is obtained in 12 min using a 400 MHz NMR spectrometer. For comparison, 2D gel cultures and 3D multicoaxial BALs were batch cultured, with medium changed every day for 15 days after inoculation with human liver cells in Matrigel-collagen type 1 gels. Glutamine consumption was higher by day 8 in the BAL than in 2D culture; lactate production was lower through the 15-day culture period. Alanine was the primary amino acid produced and tracked with lactate or urea production. Glucose and pyruvate consumption were similar in the BAL and 2D cultures. NMR analysis permits quality assurance of the bioreactor by identifying contaminants. Ethanol was observed because of a bioreactor membrane "wetting" procedure. A biochemical scheme is presented illustrating bioreactor metabolomic footprint results and demonstrating how this can be translated to modify bioreactor operational parameters or quality assurance issues.

High-Throughput Nuclear Magnetic Resonance Metabolomic Footprinting for Tissue Engineering

High-Throughput Nuclear Magnetic Resonance Metabolomic Footprinting for Tissue Engineering

High-Throughput Nuclear Magnetic Resonance Metabolomic Footprinting for Tissue Engineering

High-Throughput Nuclear Magnetic Resonance Metabolomic Footprinting for Tissue Engineering

An eight-coil high-frequency probehead design for high-throughput nuclear magnetic resonance spectroscopy

In order to increase the throughput of high-resolution nuclear magnetic resonance spectroscopy a multiple-coil probe, which enables the simultaneous analysis of eight different samples, was designed. The probe, consisting of eight identical solenoidal coils, was constructed for operation at 600 MHz. By using four receivers and radiofrequency switches, spectra from eight different chemical solutions were acquired in the time normally required for one. Two-dimensional COSY, gradient COSY, and TOCSY data have been acquired. Intercoil electrical isolation was between 25 and 45 dB, with signal cross-talk between ∼1 and 5% measured by NMR. The spectral linewidths for the eight coils were between 3 and 6 Hz for a single optimized shim setting.

High-throughput nuclear magnetic resonance analysis using a multiple coil flow probe.

An automated method for high-throughput nuclear magnetic resonance (NMR) spectroscopy has been developed using a four-coil Multiplex NMR probe. The probe is constructed with solenoidal microcoils optimized for detection of small volume, mass-limited samples and a flow-through design. Four samples can be simultaneously injected into the Multiplex probe with a robotics liquid handler and then analyzed in rapid succession using a selective excitation experiment. Due to the simultaneous injection of four samples and the reduced analysis time with rapid selective excitation, the analysis rate achieved thus far is as low as 1 sample/34 s for 1D 1H NMR.

Magnetic Resonance Diagnostics: A New Technology for High-Throughput Clinical Diagnostics

Magnetic resonance diagnostics (MRD) uses automated, high-throughput nuclear magnetic resonance (NMR) spectroscopy for the rapid identification and quantification of small-molecule metabolites in biofluid mixtures (blood, urine, saliva, cerebrospinal fluid, and others). Specifically, MRD involves using a high-field (400 MHz) NMR instrument equipped with a small-volume flow probe and robotic sample handler to rapidly load biofluid samples and to collect their 1H NMR spectra. Spectral deconvolution software automatically assigns individual peaks to particular compounds and calculates concentrations from peak areas. MRD uses the principle of chemical shift separation to physically separate and identify individual compounds directly from 1H NMR spectra, thus avoiding chromatographic separation steps (e.g., HPLC, gas chromatography, and capillary electrophoresis). MRD is useful for rapid (<2 min per sample) qualitative and quantitative assessment of small-molecule metabolites. NMR spectroscopy is not new to the field of clinical chemistry. Indeed several important applications have already been demonstrated in the area of diagnosis and therapeutic monitoring of metabolic disorders (1)(2)(3)(4), in toxicologic and renal testing (5)(6), and in the profiling of blood lipoproteins and cholesterol (7). An emerging approach to enable high-throughput in vivo toxicology is called metabonomics, which uses high-resolution NMR to rapidly evaluate the metabolic status of an animal (8)(9). A key limitation to all of these NMR approaches is that they depend on manual sample handling and/or manual (i.e., expert) spectral analysis. This has made most NMR approaches to clinical analyses far too slow or too costly for routine chemical profiling or high-throughput screening. Because MRD is fully automated (sample handling, spectral collection, and spectral analysis are all handled by robots or computers), this technique offers the potential for high-throughput, comprehensive, and inexpensive chemical analysis of a wide range of biofluid samples. To demonstrate the potential …

Multiple solenoidal microcoil probes for high-sensitivity, high-throughput nuclear magnetic resonance spectroscopy.

Two designs for incorporating multiple solenoidal microcoils into a single probe head are presented to increase the throughput of high-resolution NMR. Through a combination of radio frequency switches and low-noise amplifiers, multiple NMR spectra can be acquired in the same time as a single spectrum from a conventional probe consisting of one coil. Since this method does not compromise sensitivity with regard to the individual microcoils, throughput increases linearly with the number of coils. Only one receiver is needed, and data acquisition parameters can be optimized for each sample. Specifically, a four-coil system has been implemented for proton NMR at 250 MHz using a wide-bore magnet, with an observe volume of 28 nL for each microcoil. Signal cross-contamination was approximately 0.2% between individual coils, and simultaneous one- and two-dimensional spectra have been obtained from samples of fructose, galactose, adenosine triphosphate, and chloroquine (7 nmol of each compound). A more compact two-coil configuration has also been designed for operation at 500 MHz, with observe volumes of 5 and 31 nL for the two coils. One- and two-dimensional spectra were acquired from samples of 1-butanol (55 nmol) and ethylbenzene (250 nmol).