To obtain a comprehensive assessment of metabolite levels from extracts of leukocytes, we have recorded ultra-high-resolution 1 H-13 C HSQC NMR spectra of cell extracts, which exhibit spectral signatures of numerous small molecules. However, conventional acquisition of such spectra is time consuming and hampers measurements on multiple samples, which would be needed for statistical analysis of metabolite concentrations. Here we show that the measurement time can be dramatically reduced without loss of spectral quality when using non-linear sampling (NLS) and a new highfidelity Forward Maximum-entropy (FM) reconstruction algorithm. This FM reconstruction conserves all measured time domain data points and guesses the missing data points by an iterative process. This consists of discrete Fourier transformation of the sparse time-domain data set, computation of the spectral entropy, determination of a multidimensional entropy gradient, and calculation of new values for the missing time domain data points with a conjugate gradient approach. Since this procedure does not alter measured data points it reproduces signal intensities with high fidelity and does not suffer from a dynamic-range problem. As an example we measured a natural abundance 1 H-13 C HSQC spectrum of metabolites from granulocyte cell extracts. We show that a high-resolution 1 H-13 C HSQC spectrum with 4k complex increments recorded linearly within 3.7 days can be reconstructed from 1/7 th of the increments with nearly identical spectral appearance, indistinguishable signal intensities and comparable or even lower root mean square (rms) and peak noise patterns measured in signal-free areas. Thus, this approach allows recording of ultra-high resolution 1 H-13 C HSQC spectra in a fraction of the time needed for recording linearly sampled spectra.
In this paper we describe the development of a 5 mm NMR flow tube that can be used in a standard 5 mm NMR probe, enabling the user to conduct experiments on flowing samples or, more specifically, on flowing reaction mixtures. This enables reaction monitoring or kinetic experiments to be conducted by flowing reaction mixtures from a reaction vessel to detection in the coil area of the NMR, without the need for a specialized flow NMR probe. One of the key benefits of this flow tube is that it provides flexibility to be used across a range of available spectrometers of varying magnetic field strengths with a standard 5 mm probe setup. The applicability of this flow tube to reaction monitoring is demonstrated using the reaction of p-phenylenediamine and isobutyraldehyde to form the diimine product.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.