Chromatography is a process of separation where the analyte is stored inside a traveling liquid or gaseous medium and is pumped into a stationary stream. One step is normally hydrophilic, and the other lipophilic. The analyte components deal with those two processes differently. They spend more or less time communicating with the stationary phase based on their polarity and are thus delayed to a greater or lesser degree. Which contributes to the isolation of the different components in the sample. That sample portion elutes its retention time at a given time from the stationary phase. When the components pass through the detector their signal is reported in the form of a chromatogram and plotted.
NMR spectroscopy is one tool that can provide much of the structural information necessary and often in a non-destructive manner. NMR is useful for studying both amorphous and crystalline materials unlike X-ray diffraction spectroscopy, which requires a crystalline sample. NMR spectroscopy can be used to provide the determination of chemical purity and quantitative measurements of impurities in materials. The precision and accuracy of quantitative NMR measurements are equivalent to other forms of instrumental study.
General Principle of Chromatography
Chromatography is the name given to a particular family of separation methods where the separation is based on differences in rates of migration when the sample components are transported by a mobile phase through a stationary phase. The mobile phase can be gas, a liquid, or a supercritical fluid and the stationary phase may be a solid, a liquid or a gel. The general principle of chromatography is quite simple. Chromatography is considered the most important analytical technique in pharmaceutical analysis.
All chromatographic separations are based on a common concept; the distribution of components in a mixture between two immiscible phases, one being a stationary phase, the other a mobile phase. In order for separation of compounds to occur a dynamic or moving situation must be initiated. This is accomplished by causing one of the phases to flow past the other the stationary phase. This movement may be accomplished by way of capillary action, gravity forces or application of pressure to the mobile phase.
NMR Spectroscopy Uses
NMR spectroscopy was first introduced by Isidor Radi in 1938. Since its induction as one of the most powerful tools for structural identification, the NMR spectroscopy today has several important uses. For proteins, the NMR is used to determine the structure, locate binding sites, understand function and dynamics, and examine folding patterns of proteins. Chemists regularly use NMR spectroscopy to analyze chemical structures using simple, one-dimensional techniques. The arrangement of more complex molecules is calculated using two-dimensional techniques.
NMR spectroscopy is a versatile tool with which to determine and monitor various types of protein binding. Protein-protein and protein-ligand interactions regulate key biological processes such as signal transduction and using NMR spectroscopy is ideally suited to determine these weak interactions. Additionally NMR spectroscopy has been used with isotope editing or filtering in studies of the dynamics of protein-ligand complexes.