Principles of separation techniques

All separation procedures depend primarily upon some physical characteristic of the compounds. It is relatively easy to separate substances that have significantly different physical characteristics by simple techniques, such as solvent extraction. However, if the various compounds are similar to each other then it is essential that any slight differences between them are exploited in order to achieve separation. Table 3.2 lists the major physical properties that form the basis of separation procedures. 

It is possible to exaggerate any differences by repeating a manipulation many times, for instance, in sequential solvent extractions. Paper chromatography is an example of a separation procedure in which a large number of partition equilibria occur between an organic solvent and the water in the cellulose. 

The possible formation of a dipole is a feature of covalent bonding but it is obvious that an ionic bond results in a definite unequal distribution of electrons within a molecule and such molecules (or ions) are extremely polar. However, the fact that they carry a definite charge enables additional separation techniques to be applied. The rate of migration in an electric field (electrophoresis) and the affinity for ions of opposite charge (ion-exchange chromatography) are extremely valuable techniques in the separation of ionic species. 

Molecules that vary significantly in their size can be separated by ultrafiltration or dialysis, while molecules that are only slightly different in size can often be separated by gel permeation chromatography. Ultracentrifugal techniques, while apparently separating on the basis of size, are strictly speaking more influenced by the mass and density of the molecule and to a lesser extent by its shape. 

Many molecules show a definite affinity for another molecule based on a shape relationship, e.g. enzymes and their substrates, antibodies and their 

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Principle of separation Technique Capacity Yield Resolution Cost... 

FIG. 18-124 Operating principles of a roll-discharge mechanism. (Schweitzer, Handbook of Separation Techniques for Chemical Engineers, p, 4-40, Copyiight 1979 hy McGraw-Hill, Inc, Used with peimission of McGraw-Hill Book Co,)... 

Although is has been in use for over 50 years and has become one of the most widely used routine analysis techniques, GC research remains vibrant and challenging. There are two key areas in which dramatic advancements are being made sampling and sample introduction methods and multi-dimensional separations. A summary of sampling techniques in use with GC is shown in Table 14.9, including the basic principle of the technique and some key applications. These techniques have become critical in extending the use of GC into the diverse fields described in the Applications section. 

The classification of separation techniques as shown in Table 3.2 is concise and easy to remember but it is also simplistic because it appears to imply that only one factor is involved in each technique. In practice, the effectiveness of any method is a composite of many factors, the one indicated in the table usually being the most significant. Some of the developments in separation procedures exploit this range of factors involved in any separation technique by using conditions or reagents designed to minimize one or maximize another. As a consequence, the techniques and instrumentation of separation methods are constantly changing but the fundamental principles remain the same and need to be understood in order to appreciate the usefulness and limitations of any particular technique. 

Although the general principles of separation processes are applicable widely across the process industries, more specialised techniques are now being developed. Reference is made in Chapter 13 to the use of supercritical fluids, such as carbon dioxide, for the extraction of components from naturally produced materials in the food industry, and to the applications of aqueous two-phase systems of low interfacial tensions for the separation of the products from bioreactors, many of which will be degraded by the action of harsh organic solvents. In many cases, biochemical separations may involve separation processes of up to ten stages, possibly with each utilising a different technique. Very often, differences in both physical and chemical properties are utilised. Frequently... 

The individual variances in Equation (21) cannot be suppressed to a zero value as they are inherent to the principle of the technique. It should be possible, however, to control the contributions of these sources of variance by proper instrumental design and selection of optimal working conditions. The extent of the dispersion will affect the efficiency of the separation system, which is usually expressed in terms of the number of theoretical plates (N)... 

Ion chromatography (1C) is a separation technique related to HPLC. However, because it has so many aspects such as the principle of separation and detection methods, it requires special attention. The mobile phase is usually composed of an aqueous ionic medium and the stationary phase is a solid used to conduct ion exchange. Besides the detection modes based on absorbance and fluorescence, which are identical to those used in HPLC, ion chromatography also uses electrochemical methods based on the presence of ions in a solution. The applications of ion chromatography extend beyond the measurement of cations and anions that initially contributed to the success of the technique. One can measure organic or inorganic species as long as they are polar. 

Planar chromatography, also known as Thin Layer Chromatography (TLC), is a technique related to HPLC but with its own specificity. Although these two techniques are different experimentally, the principle of separation and the nature of the phases are the same. Due to the reproducibility of the films and concentration measurements. TLC is now a quantitative method of analysis that can be conducted on actual instruments. The development of automatic applicators and densitometers has lead to nano-TLC, a simple to use technique with a high capacity. 

NCE is a relatively new development in separation science, especially in proteomics and genomics. In the last two decades NCE has gained increasing importance, as can be seen from a good number of publications [17-20]. In addition to the above advantages, NCE is a suitable technique for samples that may be difficult to separate by NLC as the principles of separation are entirely different. Lower detection limits of NCE lead to the possibility of separating and characterizing small quantities of materials. Moreover, the enzymatic reactions for analytical purposes can be conducted within the capillary. 

The identification of the first transuranium elements was by chemical means. In the early 1960s physical techniques were developed which allowed for detection of nuclei with lifetimes of less than one second at high sensitivity. A further improvement of the physical methods was obtained with the development of recoil separators and large area position sensitive detectors. As a prime example for such instruments, we will describe the velocity filter SHIP (Separator for Heavy-Ion reaction Products) and its detector system, which were developed at the UNILAC. The principle of separation and detection techniques used in the other laboratories is comparable. 

The principles of separation of antibodies by chromatography are based on specific interactions on solid phase surfaces. Beyond their biological properties and role, antibodies possess a variety of common characteristics that can be exploited to develop a generic separation procedure. In spite of a large spectrum of existing separation techniques for antibodies, development of novel approaches is still an attractive task. Available methodologies are not applicable to all antibodies. Furthermore requirements on therapeutic antibodies are continuously increasing. 

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