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A 4D approach for protein 1H chemical shift prediction was explored. The 4th dimension is the molecular flexibility, mapped using molecular dynamics simulations. The chemical shifts were predicted with a principal component model based on atom coordinates from a database of 40 protein structures. When compared to the corresponding non-dynamic (3D) model, the 4th dimension improved prediction by 6–7%. The prediction method achieved RMS errors of 0.29 and 0.50 ppm for Hα and HN shifts, respectively. However, for individual proteins the RMS errors were 0.17–0.34 and 0.34–0.65 ppm for the Hα and HN shifts, respectively. X-ray structures gave better predictions than the corresponding NMR structures, indicating that chemical shifts contain invaluable information about local structures. The 1H chemical shift prediction tool 4DSPOT is available from . [ABSTRACT FROM AUTHOR]

A protocol for quantitative 1H NMR analysis of human cerebrospinal fluid (hCSF) was built up and assessed as based on Constrained Total-Line-Shape (CTLS) fitting. In this method, linear constraints were applied to spectral structures. The 1H NMR spectra of 45 human CSF samples were measured and quantified using the CTLS method. The quantification strategies based on total-line-shape fitting are discussed. The metabolic model for CTLS includes 31 metabolites covering 85% of the total spectral intensity, excluding the protein contribution. Prior to data analysis, the data was divided into patients with no Alzheimer’s disease (AD), but with a normal AD marker profile (the peptide β-amyloid42 and tau protein) present in CSF, and into controls that do not have an AD marker profile in CSF. Unexpectedly large variations in metabolite concentrations within the two patient groups were detected, but an analysis of variance revealed a significant ( P = 0.027) difference only in the concentration of creatinine which was higher in patients that had a normal AD marker profile. Multivariate classification tools such as self-organizing maps (SOM) failed in separation of the two classes. [ABSTRACT FROM AUTHOR]

This study introduces a new NMR-based methodology for identification (ID) and quantification (purity, strength) assays of widely used amino acids. A detailed analysis of four amino acids and their available salts was performed with both a high-field (600 MHz) and a benchtop (60 MHz) NMR instrument. To assess sensitivity constraints, samples for 1H NMR analysis were initially prepared using only 10 mg of analyte and 1 mg of maleic acid (MA) as an internal calibrant (IC) and secondary chemical shift reference. The characteristic dispersion of the peak patterns indicating the presence or absence of a counterion (mostly chloride) was conserved at both high and low-field strength instruments, showing that the underlying NMR spectroscopic parameters, i.e., chemical shifts and coupling constants, are independent of the magnetic field strength. However, as the verbal descriptions of 1H NMR spectra are challenging in the context of reference materials and pharmaceutical monographs, an alternative method for the identification (ID) of amino acids is proposed that uses 13C NMR patterns from multiplicity-edited HSQC (ed-HSQC), which are both compound-specific and straightforward to document. For ed-HSQC measurements, the sample amount was increased to 30 mg of the analyte and several acquisition parameters were tested, including t 1 increments used in the pulse program, number of scans, and repetition time. Excellent congruence with deviations <0.1 ppm was achieved for the 13C chemical shifts from 1D 13C NMR spectra (150 MHz) vs. those extracted from ed-HSQC (15 MHz traces). Finally, all samples of amino acid candidate reference materials were quantified by 1H qNMR (abs-qHNMR) at both 600 and 60 MHz. At high field, both IC and relative quantitations were performed, however, with the low-field instrument, only the IC method was used. The results showed that the analyzed reference material candidates were generally highly pure compounds. To achieve adequately low levels of uncertainty for such high-purity materials, the sample amounts were increased to 100 mg of analytes and 10 mg of the IC and replicates were analyzed for selected amino acids. [Display omitted] • 1H NMR identifies free amino acids and distinguishes them from their salts. • Indirect 13 C NMR measurement via 2D HSQC can serve as innovative identity assay. • Congruence in IC- and rel-qHNMR is diagnostic for chemical purity and water content. • Sample size and replicates are key for absolute qHNMR or high-purity compounds. • Benchtop NMR is suitable for pharmaceutical identity, quantity, and purity assays. [ABSTRACT FROM AUTHOR]

The characteristic signals observed in NMR spectra encode essential information on the structure of small molecules. However, extracting all of this information from complex signal patterns is not trivial. This report demonstrates how computer-aided spectral analysis enables the complete interpretation of 1D (1)H NMR data. The effectiveness of this approach is illustrated with a set of organic molecules, for which replicas of their (1)H NMR spectra were generated. The potential impact of this methodology on organic chemistry research is discussed.