Two-Dimensional NMR Spectroscopy: Background and Overview of the Experiments [New Analytical Methods (36)]

An exact knowledge of the structure, dynamics, and reactions of molecules provides the key to understand their functions and properties. NMR spectroscopy has developed, through the introduction of two-dimensional methods, into the most important method for the investigation of these questions in solution. A great variety of different techniques is available. However, for their successful application not only the appropriate equipment is required, but also the right choice of experiments and optimum measurement parameters, as well as a careful evaluation of the spectra. This contribution describes the necessary background for modern NMR spectroscopy. With the aid of the so-called product operator formalism it is possible to understand pulsed Fourier transform NMR spectroscopy both qualitatively and quantitatively. Very few, readily understandable assumptions are sufficient for confident application of these methods. This article attempts to introduce in a simple manner this formalism as well as phase cycles necessary for the understanding of pulse sequences, and to train the reader through the discussion of several 2D NMR techniques. An overview of the most important techniques is given in the second part of this article.     

Two-Dimensional Solid-State NMR Spectroscopy: New Possibilities for the Investigation of the Structure and Dynamics of Solid Polymers [New Analytical Methods (38)]

NMR spectroscopy is an effective method not only for examining liquid samples but also for characterizing molecular sturcture, order and dynamics in amorphous and ordered solids. Recent developments in the area of solid-state NMR spectroscopy span from model-dependent studies of conventional one-dimensional spectra to the more definitive two-dimensional (2D) spectra which provide more specific information. For example, with 2D-NMR spectroscopy it is possible to determine the orientational distribution functions of molecular segments in drawn polymers and to distinguish different mechanisms of complex molecular motions. Following an introduction to basic NMR spectroscopy, an overview of the current state-of-the-art of 2D methods in solid-state NMR spectroscopy is presented and demonstrated with selected examples.     

Chromatographic Resolution of Racemates. New analytical methods (17)

A number of racemates can be resolved into optically pure enantiomers by chromatography on optically active adsorbents. Synthetic polymers with optically active amide, amino acid, and crown ether groups, natural products such as starch and cellulose, and also microcrystalline triacetylcellulose are suitable for this purpose. Racemates can also be resolved by gas chromatography on optically active stationary phases.     

High-Resolution Metal NMR Spectroscopy of Organometallic Compounds [New Analytical Methods (30)]

Great advances have been made in the past decade in the field of NMR spectroscopy. Apart from the development of completely new areas of application, such as in solid-state chemistry, in materials science, in physiological chemistry, and in medicine, with the introduction of new pulse spectroscopic methods and the application of high magnetic field strengths important progress has also been made in the traditional field of high-resolution NMR spectroscopy. Thus, among other things, the observation of metal resonances has been facilitated and new areas of application have been opened up in inorganic and organometallic chemistry. In this review, recent detection methods for spin-1/2 and quadrupolar metal nuclei are presented and discussed. The use of metal-NMR spectroscopy with respect to problems of a typical chemical nature, mainly from the field of organometallics, is demonstrated for a number of selected metal nuclei (²⁵Mg, ²⁷Al, ⁴⁹Ti, ⁵⁷Fe, ⁵⁹Co, ⁶¹Ni, ⁹¹Zr, ¹⁰³Rh, and ¹⁹⁵Pt). Relations found empirically between chemical shifts and coordination number, oxidation number, and electronic configuration of a metal bound in a complex are emphasized. Furthermore, cases in which the chemical shifts of metal nuclei can be interpreted in terms of the energy difference of frontier orbitals are presented. This aspect leads to the establishment of a relationship between chemical reactivity and NMR parameters for a series of related compounds.     

The Electro-Optical Kerr Effect in Conformational Analysis. New analytical methods (10)

Birefringence induced in a material by application of an electric field is known as the Kerr effect. The great sensitivity of this property to variations of molecular geometry and environment has made it the basis of a potent method for studying the conformations of molecules. It has, in addition, been effectively applied to investigations of the geometry of solvation and of solute aggregation. An outline is presented of the general technique of measurement of the Kerr effect, the extraction of directed effective polarizabilities of solute molecules and of structural groups, the calculation of theoretical Kerr constants for candidate structures of molecules, and the procedures used to determine solute conformations. The scope and limitations of the method are illustrated by specific conformational problems drawn from the literature. In the case of highly flexible molecules analysis of the measured Kerr constant is inhibited by uncertainties in conformational energy data. This paper is concerned mainly with the specification of detailed conformational energy data. This paper is concerned mainly with the specification of detailed conformational features of molecules. Reference is made, however, to the utilization of pulsed orienting fields to study shapes and dimensions of biological macromolecules.     

15N-NMR Spectroscopy—New Methods and Applications [New Analytical Methods (28)]

The nitrogen nucleus is the third most important probe (after ¹H and ¹³C) for structural investigations of organic and bioorganic molecules by NMR spectroscopy. For a long time, however, the insufficient sensitivity and low natural abundance of the ¹⁵N isotope hampered detection of the ¹⁵N nucleus, and the quadrupolar ¹⁴N nucleus proved unsuitable for the study of larger molecules with several nonequivalent nitrogen atoms. The advent of new techniques, such as pulse sequences and polarization transfer, in conjunction with the use of high-field magnets and large-sample probe heads largely solved the detection problem. As a result, the last few years have seen a dramatic development of ¹⁵N-NMR spectroscopy as a versatile method for studying molecular structure, both in isotropic (liquid) and anisotropic (solid) phases. The scope of chemical applications extends from inorganic, organometallic, and organic chemistry to biochemistry and molecular biology, and includes the study of reactive intermediates, biopolymers, enzyme-inhibitor complexes, and nitrogen metabolism. Two-dimensional NMR techniques offer additional possibilities for detailed studies of biological systems.     

Physicochemical Principles and Applications of Supercritical Fluid Chromatography (SFC). New analytical methods (19)

In supercritical fluid chromatography (SFC) compressed gases in the region of their critical temperature are used as mobile phases. SFC has important advantages over gas chromatography (GC) for the separation of low-volatile or thermally unstable substances. Like high pressure liquid chromatography (HPLC) and gel chromatography, it is used for various special applications and preparative separations, e.g. in the petroleum industry and in the separation of oligomers. SFC is of great interest in fundamental research on fluid extraction and for the determination of the physicochemical properties of fluid systems. In this contribution the most important physicochemical, methodological, and instrumental principles of SFC are summarized; characteristic physicochemical applications are the determination of capacity ratios, partition coefficients, partial molar volumes, interaction second virial coefficients, and difusion coefficients.     

High-Resolution Solid-State 13C-NMR Spectroscopy of Polymers [New Analytical Methods (37)]

Until a few years ago, solid-state nuclear resonance yielded spectra containing broad lines only. Meanwhile, CP/MAS-NMR spectroscopy has provided a method which gives narrow nuclear resonance lines from a solid-state specimen as well. Using this technique, it is now possible to produce spectra of “rare” nuclei (¹³C, ²⁹Si, ¹⁵N etc.) which are resolved in terms of chemical structure. The analytical capabilities of NMR spectroscopy can be applied to the solid state: it may be that it is necessary to identify compounds in the solid state because, for example, a solvent would alter the coordination sphere, or that it is desired to monitor chemical reactions in the solid state, for example the baking of an enamel. Where a substance in the solid state is concerned, high-resolution ¹³C-NMR spectroscopy provides not only information about the chemical structure, but also about the solid state itself. To mention just a few examples, information on the conformation, crystal structure and molecular dynamics, as well as molecular miscibility is given. This opens up a broad spectrum of applications, from a statement concerning the crystal modification of an active substance in ready-to-use pharmaceutical preparations, e.g. tablets, to the question of whether two polymers are miscible with one another at a molecular level.     

Fully Automatic Potentiometric Titrations [New analytical methods (9)]

Fully automatic titration systems consist of at least two units: the sample-changing system and the measuring system, the latter including addition of the reagent. These systems are preferably coupled to a computer. The possibilities of fully automatic titration can be best utilized if the sources of error are taken into account and the evaluation procedure optimal for the specific analysis is selected.     

NMR Microscopy—Fundamentals,Limits and Possible Applications

Alongside the numerous applications of NMR spectroscopy to structural elucidation in analytical chemistry, and to biochemical and morphological studies by NMR tomography, NMR microscopy makes possible a whole new range of applications. These include imaging, the investigation of biological objects such as plants and small animals, and also the observation of microscopic structures and structural changes in polymers and ceramics. NMR spectroscopy can also be conducted combinationally as volume-selective spectroscopy, whereby it is possible to spatially resolve the NMR-specific parameters: spin density ?, chemical shifts δ, and the relaxation times T₁ and T₂. The numerous well developed methods available make it possible to study dynamic processes by fast imaging with a temporal resolution in milliseconds. This not only allows the imaging of moving objects without incurring movement artefacts but also the measurement of diffusion constants in isotropic and anisotropic diffusion—in the latter case allowing, in principle, the determination of the complete diffusion tensor. The spatially resolved measurement of the relaxation times yields information on molecular mobility and bonding, e. g. the bonding of water, or other solvents, to polymers, the mobility of fluids in polymers or ceramics, or the three-dimensional evaluation of pore size in porous materials. In biomedicine, NMR microscopy allows the observation of growth on the cellular level, the study of embryos, and the development of therapeutic methods in animal experiments. It can lead to a drastic reduction in the number of animal experiments, and in combination with volume-selective spectroscopy gives valuable information on in-vivo metabolism.     

State of Development of Atomic Absorption Spectrometry [New analytical methods (1)]

Atomic absorption spectrometry has developed extremely rapidly in recent years, and is now used in many analytical laboratories. The purpose of this progress report is to show the present position and to examine critically the possibilities and limitations of atomic absorption methods.     

Highly Resolved Pure-Shift Spectra on a Compact NMR Spectrometer

Benchtop NMR spectrometers experience a great success for a wide range of applications. However, their performance is highly limited by peak overlaps. Emerging “pure-shift NMR” (PS NMR) methods have been intensively used at high field to enhance the resolution by homodecoupling strategies. Here, different PS methods have been implemented on a compact NMR spectrometer operating at 43 MHz. Among the PS methods, the recent PSYCHE scheme appears more sensitive than Zangger-Sterk (ZS) experiments and offers a substantial resolution improvement as compared to 1D ¹H. On the other hand, despite their slightly lower sensitivity, ZS methods are more efficient to reduce broad signals and are more immune to strong couplings. Finally, the classical J-resolved pulse sequence is more efficient to reduce larger signals for bigger-sized molecules. The three approaches appear relevant for benchtop NMR and their combination forms an efficient toolbox to analyze a great diversity of samples.     

Resonance Raman Spectroscopy,and Its Application to Inorganic Chemistry. New Analytical Methods (27)

Resonance Raman spectra are obtained when the wave number of the exciting radiation is close to, or coincident with, that of an electronic transition of the scattering species. Such spectra are usually characterized by a very large enhancement of the intensities of particular Raman bands, sometimes with the appearance of intense overtone and combination tone progressions. The technique provides detailed information about excited electronic states because it is only the vibrational modes associated with the chromophore that are resonance-Raman active. Additionally, the high sensitivity is such that compounds at concentrations as low as 10^?6 mol/L may be detected, enabling resonance Raman spectroscopy to be used as an analytical tool and for the study of chromophores in molecules of biological interest.     

Direct Identification of the Carbon Skeleton of Organic Compounds using Double Quantum Coherence 13C-NMR Spectroscopy. The INADEQUATE Pulse Sequence

NMR spectroscopy plays an important part in the determination of the structures of organic compounds. The parameters of importance here are the chemical shifts of the ¹H and ¹³C nuclei and the spin-spin interactions both between ¹H nuclei and between ¹H and ¹³C nuclei. Couplings between ¹³C nuclei were almost completely neglected until a few years ago, since they were extremely difficult to observe because of the low natural abundance of ¹³C. However, it is these couplings which afford information directly on the carbon-carbon connectivities in the molecule. It is now possible to use a special NMR pulse sequence to make these couplings more readily visible: the result of using this sequence is a ¹³C-NMR spectrum from which the carbon skeleton concerned can be directly read off. Two-dimensional spectra in particular are very easy to evaluate. The pulse sequence involved, which bears the somewhat puzzling name INADEQUATE, produces double-quantum coherences from which the NMR signals of the coupled carbon nuclei can be obtained. In this article the principle of double-quantum coherence is described and a number of examples for the application of the INADEQUATE pulse sequences to problems in synthetic organic chemistry, biosynthesis and natural products chemistry are presented; in addition, the possibility of applying the INADEQUATE method to other nuclei is considered.     

Validation and qualification: the fitness for purpose of mass spectrometry-based analytical methods and analytical systems

The context of validation for mass spectrometry (MS)-based methods is critically analysed. The focus is on the fitness for purpose depending on the task of the method. Information is given on commonly accepted procedures for the implementation and acceptance of analytical methods as ‘confirmatory methods’ according to EU criteria, and strategies for measurement. Attention is paid to the problem of matrix effects in the case of liquid chromatography-mass spectrometry-based procedures, since according to recent guidelines for bioanalytical method validations, there is a need to evaluate matrix effects during development and validation of LC-MS methods “to ensure that precision, selectivity and sensitivity will not be compromised”. Beneficial aspects of the qualification process to ensure the suitability of the MS analytical system are also evaluated and discussed.     

Cyclic Voltammetry—“Electrochemical Spectroscopy”. New Analytical Methods (25)

Polarography is still the best known classical measuring method in electroanalytical chemistry. However, in recent years its position has been challenged by cyclic voltammetry (CV). Simple diagnostic criteria and relatively easily acquired measuring techniques have hastened this development. Cyclic voltammetry has the further attraction of providing information not only on the thermodynamics of redox processes but also on the kinetics of heterogeneous electron-transfer reactions and coupled chemical reactions. The characteristic shapes of the voltammetric waves and their unequivocal position on the potential scale virtually fingerprint the individual electrochemical properties of redox systems. For this reason the method has been labeled “electrochemical spectroscopy”.     

The Aza‐Morita–Baylis–Hillman Reaction: A Mechanistic and Kinetic Study

The aza‐Morita‐Baylis–Hillman (aza‐MBH) reaction has been studied in a variety of solvents, a selection of imine substrates and with various combinations of PPh₃ and para‐nitrophenol as the catalyst system. The measured kinetic data indicates that the effects of solvent and protic co‐catalyst are strongly interdependent. These results are most easily reconciled with a mechanistic model involving the reversible protonation of zwitterionic intermediates in the catalytic cycle, which is also supported by ³¹P NMR spectroscopy and quantum chemical studies.     

(14)N NMR and two-dimensional suspension (1)H and (13)C HRMAS NMR spectroscopy of ionic liquids immobilized on silica

A variety of popular ionic liquids have been synthesized and characterized, including by optimized (14)N NMR spectroscopy of the neat and dissolved ionic liquids. Ionic liquids incorporating Si(OEt)(3) groups have been immobilized on silica in a well-defined manner with the imidazolium moiety remaining intact. This has been proved by optimized one- and two-dimensional (1)H and (13)C HRMAS NMR spectroscopy of the materials suspended in suitable solvents.     

Liquid Column Chromatography with Chemical Derivatizations after Separation. [New analytical methods (15)]

Modern liquid column chromatography (high-pressure liquid chromatography, HPLC) has evolved in the last few years into a highly efficient and versatile separation technique. The selectivity of an analytical process that depends upon a previous separation step can in many cases be increased considerably by chemical derivatizations after the separation. In addition, lower detection limits can be achieved in this way than in detection without derivatization. The physicochemical principles of these combined processes involving chromatographic separation and chemical derivatization prior to detection (coupling of HPLC and a reaction detector) are presented and discussed. The state of development is outlined, with a survey of the more important applications so far described in the literature.