Case studiesExamples of different NMR case studies performed at the NMR expertise centre
Case studies
Below a number of studies are summarized where NMR played a critical role in the elucidation of the final structures, corresponding synthesis pathways, molecule properties, etc. For more detailed information, please see the corresponding article linked at the bottom of each example.
NMR in Metabolites
Structure elucidation and confirmation of various small to medium sized plant metabolites
Given a 3-OGlc-echinocystic acid sample limited in quantity (< 1mg) and questionable purity the following questions needed to be answered:
What is the position of hydroxylation in the 3-O-Glc-echinocystic acid metabolite and what is it's relative stereochemistry?
Can the overall structure of the metabolite be confirmed?
No additional or unexpected functionalities present?
Through a combination of 1D 1H and 13C spectra (left) for comparison against a known 3-Glc-Echinocystic acid standard and 2D 1H-1H, 1H-13C spectra including HSQC and HMBC-type experiments (right) the structural integrity of the metabolite was proven to be identical. In addition, where initially the additional -OH group was expected to be located at position 21, it was shown to be present at position 16.
The spectra shown were recorded on 700MHz with the use of a shigemi tube in order to boost concentration.
In collaboration with Alain Goossens - Flemish Institute for Biotechnology (VIB)
Structure elucidation and confirmation of peptides, peptide modifications and peptide by-products
Given a proof-of-principle peptide sequence in a reversible/irreversible and orthogonal peptide labeling strategy, can the following questions be answered:
Can the sequence be assigned and proven to be intact following the labeling reaction?
What is the structure of the furan based peptide label given the two most likely possibilities (A or B)?
Following the classical peptide assignment methods (2D 1H-1H TOCSY and NOESY measurements @ 500MHz), additional ROESY measurements were performed at -50°C in order to slow down the exchange of the labile -NH proton (18) and show the decisive through-space contact to the other proton involved (8). In order to make this possible, a dedicated liquid nitrogen evaporation setup was used to cool down the sample inside the probe.
hr-MAS as a tool to study polymer networks at the interface of solid materials
hr-MAS or high resolution Magic Angle Spinning is an NMR technique with a specialized setup to study materials at the interface of solution and solid state.
A sample (e.g. solvent swollen polymer sample) is spun at high frequency under the 'magic angle' (54,7°)
This allows the application of most high-resolution solution state techniques to characterize molecules grafted onto insoluble supports (e.g. interfaces, networks or other matrices)
hr-MAS NMR used to monitor and quantify chemical modifications of cryogels
In this study, hr-MAS NMR was used to study the chemical modification reactions of alkyne containing polyHEMA-based macroporous network structures (cryogels). Here, Cu(I) catalyzed azide-alkyne 'click' cycloaddtion reactions are used to chemically modify the cryogel structure. These are monitored at each step by means of NMR.
Complete conversion is obtained when benzylazide is reacted with the grafted alkyne function, but only partial conversion is observed when using the PEO-N3 i.e. azide modified poly(ethylene oxide). Addition of benzylazide in a second step further consumes the remaining alkyne groups on the cryogel backbone.
A synergy between NMR and computer assisted model building allows to establish the 3D structure of (bio)molecules
NMR techniques can be used to obtain information concerning the structure and dynamics of the samples in question. By subsequently using this information as structural restraints in modeling software such as force fields (AMBER, CNS, ...) an ensemble of 3D models that correspond with the solution structure can be obtained. The insights obtained from these models, allows for a better understanding of the function and/or properties of the sample in question.
Structure elucidation and confirmation of bio-engineered surfactants
In the design of an efficient microbial production process for amphiphilic surfactants, the structure of the resulting product needed to be characterized in order to gain insight in the overall synthesis pathway operated by the host organism. Where the main building blocks of the product were known, the following items needed to be addressed;
How is the aliphatic tail attached to the sugar entity 'A, B'?
How is sugar entity 'C, D' attached to the main part of the molecule and what is the mutual sugar connectivity?
What is the configuration of the double bond in the aliphatic tail?
What are the anomeric configurations of the four sugars?
Following an in-depth NMR analysis using 2D 1H-1H and 1H-13C spectra, the following structure was proposed.
