Applications of NMR Spectroscopy

Nuclear magnetic resonance (NMR) spectroscopy is capable of quantifying molecules. The term so called quantitative NMR (qNMR), has been used for determination of the concentration and purity of small molecules. Carbohydrates are found in various beverages and dietary foods, including crops, milk, fruits, and vegetables. Commercial products frequently use “added sugar” in soft drinks, cookies, candies, and foods. The added sugar in beverages can be sucrose, high-fructose corn syrup (HFCS) and glucose. Here, we report a quantitative method to measure 6 common sugar ingredients in foods from a single one-dimensional 1H-NMR and by using naphthimidazole (NAIM) derived sugars, which are chemically tagging aldoses with 2,3-naphthalenediamine (NADA) at the reducing ends to assist assignment of sugars. The aldoses in native sugars contain α and β anomeric isomers, and may have overlapping signals in 1H-NMR spectra. In contrast, both the anomeric isomers can be converted into a single sugar-NAIM derivative, which resolves the problem of overlapping signals to simplify the NMR quantitative analysis. This NAIM method is especially useful for identification and quantification of multiple kinds of sugars in beverages and foods. This study is to facilitate the quantification of six common sugars in beverages and foods. Our results suggest that a simple treatment of beverage and food with the NAIM labeling method provides a more extensive success rate for the quantification of sugar ingredients.

This chapter presents the correlation between coffee compounds identified by 1H Nuclear Magnetic Resonance (1H NMR) spectroscopy with psychological attention tests in order to verify which compounds are related to the focus and/or diffuse attention. Psychometric tests applied by a clinical psychologist, before and after coffee intake, were the focus attention AC-vector and the diffuse attention TADIM, and the focus attention TACOM-B and the diffuse attention TEDIF, respectively. Different tests to measure the attention before and after coffee intake were used to avoid learning effects. After AC-vector and TADIM tests, each volunteer consumed a total of 40 mL of coffee with different cup qualities (four different coffee blends – 10 mL per beverage) and indicated the order of preference in relation to the smell. This approach was used to create a greater metabolic variation between the samples tested, allowing to build a robust chemometric model. For each preferred coffee, a 1H NMR spectrum was obtained and a chemometric data treatment based on Partial Least Squares (PLS) regression and Variable Importance in Projection scores (VIP scores) was used to correlate the spectra with the psychological test results and to verify which metabolites of the coffee beverage could be related to the focus or diffuse attention. In general, our results showed that coffee intake attenuated diffuse attention and improved focus attention in most volunteers. The major metabolites that contributed to both diffuse and focus attention were caffeine, trigonelline, chlorogenic acids, acetate, lipids, lactate, γ-quinide, and polysaccharides. Among metabolites exclusively important to focus attention, formate, choline, myo-inositol, citrate, and malate were the most important. Therefore, the 1H NMR profile, in combination with chemometric tools, is interesting to assess the correlation between coffee compounds and human attention.

Nucleic acid aptamers are single-stranded DNA or RNA molecules that can fold into unique conformations and specifically recognize various targets, such as small molecules, proteins, cells, and tissues. Aptamers are selected in vitro through an iterative process called Systematic Evolution of Ligands by Exponential Enrichment (SELEX). As aptamers possess several advantages over antibodies, several diagnostic and therapeutic applications have emerged in recent years. Aptamers also attract interest as they form the receptor domain of RNA-based riboswitches that function as natural modulators of gene expression. Aptamer domain of riboswitch can sense the metabolite and this binding event is transduced into a conformational change, thereby transcriptional or translational control is achieved. Riboswitch engineering has gained importance due to the potential use of artificial riboswitches in biosensors and nextgeneration therapeutics. Therefore, understanding the structural basis of ligand binding and conformational change is critical for the success of optimization or re-engineering of aptamers. Since crystallization of aptamer-small molecule target complexes is particularly difficult, NMR provides an indispensable tool for structural analysis. In this chapter, we first give a brief information about aptamers and riboswitches. Then, we review the NMR structures of aptamers and riboswitches reported to date. We highlight the importance of NMR for identification of ligand binding mechanism, post- SELEX optimization of aptamers, as well as for the design of artificial riboswitches. In this context, we also give some examples of aptamer studies involving a combination of NMR and other techniques.

Nuclear magnetic resonance (NMR) is a special branch of spectroscopy which exploits the magnetic properties of atomic nuclei for molecular elucidation and identification. A technique that was initially developed to analyze chemical and physical molecular structure is now widely used in medical diagnosis. The noninvasiveness, non-destructiveness and simplicity of sample preparation make NMR the preferred technique for metabolomics study. Various body fluids such as urine, saliva, blood, plasma, serum and sweat have been explored to identify potential biomarkers of diseases. Psychiatric disorders, specifically alcohol-use disorder and neurological disorders such as Parkinson’s disease, have been investigated with the aid of NMR spectroscopy. Cancer has been one of the most widely studied areas and the research also includes determination of biomarkers which not only could detect the presence of cancer but also potentially predict the various cancer processes in cancer cell lines. Infectious diseases including the compounds produced by the microorganisms such as in tuberculosis and pneumonia have also been explored. Besides, NMR metabolomics has also been used to establish a metabolic fingerprint for risk stratification and early detection of cardiovascular disease (CVD). The samples of subjects with the diseases were collected and the metabolites were compared against controls such as healthy individuals using complex chemometrics and multivariate data analysis such as principal component analysis, partial least square and orthogonal partial least square analyses to distinguish the potential biomarkers. In terms of the various uses of NMR metabolomics in the subject of diagnostic medicine, more improvements to overcome the analytical limitations are expected, making it one of the most notable diagnostic tools of the future. This chapter reviewed some of the published articles in cancer, psychiatric and neurological diseases to provide examples of using NMR spectroscopy in diagnosing human disorders.

Cancer is a category of diseases characterized by uncontrolled cell growth and high potential to disseminate to other parts of the body. Cancer diagnosis is challenging due to the high structure similarity between normal and cancerous cells and the aggressive diagnostic procedures. Early diagnosis of cancer is crucial to increase the remission probability and avoid complications. A number of techniques have been involved in cancer diagnosis including biopsy, laboratory tests, computerized tomography (CT) scan, Ultrasonography, X-ray imaging, and nuclear magnetic resonance (NMR) spectroscopy. NMR has been applied both in vivo (known as magnetic resonance imaging) and in vitro to aid in cancer diagnosis. This chapter discusses the application of in vitro NMR in diagnosis and prognosis of different types of cancer with emphasis on the metabolic alterations at early stages of malignancy. The signature metabolites of brain, breast, epithelial ovarian, prostate, lung, colorectal, bladder, and oral cancers have been presented. A perspective overview of the role of NMR spectroscopy in cancer diagnosis has also been presented. This chapter shed the light on the important role of NMR spectroscopy in cancer diagnosis and treatment follow up. The applications introduced are not meant to provide a complete list of existing studies, but to present a wide overview of the current progress in this field. The chapter will cover the following topics.

Proteins are vital players that mediate a vast majority of cellular functions. NMR spectroscopy originally developed by physicists for investigation of nuclear properties, now represents highest applications in chemistry and biochemistry. NMR has been extensively utilized by structural biologists for exploring protein-ligand interactions and by medicinal chemists for drug discovery. The ligands investigated involved small organic molecules, peptides, proteins and nucleic acids. Recently, there has been increasing interest in the dynamic studies of these protein-ligand interactions. These applications are provided by a multitude of NMR experiments ranging from the simple one-dimensional 1H spectrum to complex multidimensional NMR approaches. Chemical shift perturbation analysis allows for delineation of the binding interface, determination of the dissociation constants and estimation of ligand binding kinetics. Paramagnetic Relaxation Enhancement NMR spectroscopy has been widely used to visualize the weakly populated states and describes the process of protein complex formation. These approaches have been demonstrated for substrate binding, allostery, state equilibria and macromolecular self-association. NMR spectroscopy allows for characterization of minor conformational dynamic differences in structurally similar proteins. Target Immobilized NMR screening represents another approach to drug discovery that allows ligand screening for challenging targets. NMR spectroscopy can also be applied in combination with other techniques including X-ray crystallography and various computational methods to achieve greater coverage than any of the individual methods. This chapter is focused on the applications of NMR in exploring protein-ligand interactions and dynamics.