The use of two-dimensional 1H NMR has been used to overcome problems of spectral overlap. Various applications of 1H and 13C NMR for the profiling of natural products are described.

Although 1H NMR is more sensitive, 13C NMR spectra are simpler with less overlapping signals and are less affected by different magnetic field strengths. This paper will describe the basic theoretical principles that should be considered to develop NMR into a standard quantitative method.

These approaches, however, are not mutually exclusive and will likely converge in the future. There are two general approaches to the use of NMR for profiling studies: an untargeted approach, which uses chemometric analysis and a targeted approach, which aims to quantify known compounds in the extract. Although MS is gaining popularity in metabolomics, NMR enjoys a number of key advantages because it is nondestructive, unbiased, quantitative, does not require separation or derivatization, and is amenable to compounds that are difficult to analyze by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). The last remaining peak at 4.999 ppm must be proton 13 this is confirmed by COSY correlation with proton 12, triplet multiplicity based on splitting by proton 12, and integration of one proton.Nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) are the principal methods of metabolomics, the branch of вЂ-omicsвЂ™ that deals with small molecules. We can assign proton 12 (3.564 ppm) based on its integration of 2H and its COSY correlation to proton 11. Proton 7вЂ™s peak at 6.163 ppm is split into a triplet by the two 8 protons, confirming the assignment.ДЄll that remains are protons 12 and 13. Once proton 8 has been assigned, we can easily assign proton 7 based on the remaining COSY correlation for proton 8. Therefore, we can assign proton 10 as 5.209 ppm and proton 11 as 3.754 ppm. To differentiate protons 10 and 11, take a look at our COSY table 3.754 ppm shows two COSY correlations, while 5.209 ppm only shows one. From this list, we can easily assign proton 8 as the peak at 2.068 ppm based on its integration of 2 protons. Based on the COSY, proton 9 couples protons at 2.068 ppm (2H), 3.754 ppm (1H), and 5.209 ppm (1H). ThymidineвЂ™s structure suggests that proton 9 should couple protons 8, 10, and 11. Now that proton 9 has been assigned, the fun really begins. The remaining protons are doublets, triplets, and multiplets that can be assigned by 2-dimensional COSY. The peak also integrates to 1 proton, supporting the assignment. The chemical shift of 11.256 ppm supports this assignment, as imide protons often show up far downfield. The only proton that should show up as a singlet is proton 6, as it has no neighboring protons that would split the peak (the nearest proton is 5 bonds away!). There is only one singlet in the В№H-NMR spectrum.

Therefore, the peak at 7.690 ppm must represent proton 4! The integration and chemical shift support the assignment, as proton 4 is the only aromatic proton in the structure. The long-range coupling constant observed for proton 3 (J=1.2 Hz, split into a doublet by proton 4) is reflected in the coupling constant for proton 4 (J=1.2 Hz, split into a quartet by proton 3). Protons that are coupled to each other should exhibit the same coupling constant. The peak is split into a doublet with a coupling constant of 1.2 Hz, reflecting the long-range coupling between protons 3 and 4, which also supports this assignment. The high field chemical shift supports this assignment. The only peak with an integration of 3 is the doublet at 1.770 ppm. Proton 3 is the only methyl group in the structure, and therefore must integrate to 3 protons.

Mestrenova for mass spec steps