Chapter 4.5. Ion exchange chromatography

Abstract

Publisher Summary This chapter presents a summary of the literature on ion exchange chromatography in biochemistry. Ion exchange in the form of batch processes and chromatography represents one of the most important separation methods in biochemistry. The chief advantage of this technique, compared with others—for example, gel-permeation (size-exclusion), partition and affinity chromatography and electrophoretic column methods, is much higher sorption or separation capacity. This permits considerably higher loads to be applied to columns of equal bed size. The separation possibilities are multiplied by the fact that gradients of both ionic strength and pH can be used. Classic liquid column chromatography of biopolymers (especially of proteins and nucleic acids) used to be problematic because suitable chromatographic supports were not available. Newer approaches to the development of ion exchange separation methods in biochemistry are represented by two general trends: (1) medium and high pressure liquid chromatography (MPLC, HPLC), (2) chromatofocusing. The main difference between MPLC and HPLC is not only in height of the performance, speed of the separation process, and the pressure required, but also in the instrumentation. A homemade MPLC apparatus can be improvised in nearly every laboratory using—for example, spare parts of amino acid analyzers or sugar analyzers. For the HPLC methods it is usually necessary to use commercial equipment or parts, because the requirements of construction are much more rigid. For the separation of low and middle molecular weight substances HPLC methods are mostly used.