Separation Methods in Biochemistry [antikvár]

CHAPTER 1 INTRODUCTORY Methods for the separation and isolation of the components of a mixture have been practised from the beginnings of chemical science, and the progress of chemistry has to a very great extent been bound up with the development of such techniques. The simplest group of these methods involves the separation of phases, as in filtration or the removal of a liquid phase by drying. Modern developments of these methods have been concerned mainly with technical improvements, and have added little of fundamental importance to our knowledge of separation processes. Phase separations of this type will not be considered further in this book, especially as excellent accounts of them are available elsewhere.^ Separations of a more refined nature have, however, also been used for a very long time. Crystallization and distillation were used extensively in medieval times, and they remained the chief separation methods at the disposal of the chemist until the beginning of the present century. The growth of biochemistry and the special problems and restrictions imposed by the properties of living matter have, to a very large extent, provided the impetus for the development of the wide range of separation methods available today. The study of the metabolic interrelationships in biological systems requires analytical methods of extreme sensitivity and specificity. The development of such methods is rendered diificult by a number of considerations. In the first place, the material studied is highly complex, often containing several hundreds of substances, many of which are of unknown nature. In the second place, the constituent which is to be estimated may be present in extremely small amounts, so that very high sensitivity is of paramount importance. In the third place, the systems studied are frequently very labile, so that special care is necessary to prevent the loss or modification of the components during the separation. This circumstance often necessitates working with substances in solution, at relatively low temperatures, avoiding extremes of pH and salt concentration. Again, the biochemist is often concerned with the analysis of classes of compounds of essentially the same chemical type, where the only reaction available for the final quantitative determination involves a chemical grouping common to the whole class. Examples of this kind are provided by the amino acids and the monosaccharides. In such cases, complete separation of the individual members from each other is essential before analytical determination. Another type of problem is the demonstration of homogeneity of a substance isolated from natural sources. The purity of a chemical compound is essentially a negative concept. It