Chromatography, of biomolecules

S. Fulton and D. Vanderburgh, The Busy Researcher s Guide to Biomolecule Chromatography (1996), PerSeptive Biosystems (Framingham, MA). Practical guide to chromatography of biomolecules with emphasis on perfusion methods. 

Wisniewski, R., Boschetti, E., and Jungbauer, A. (1996). Process design considerations for large scale chromatography of biomolecules Biotechnol. Biopharma. Manuf. Process. Preserv. 2, 61-181. 

In preparative chromatography, we deal with favorable adsorption isotherms, which means that the concentration in the stationary phase does not increase at the same rate as does the concentration in the mobile phase. The Langmuir adsorption isotherm originally described for adsorption of gases on plane surfaces [43] has frequently been used to describe phase equilibrium in chromatography of biomolecules [44]. 

Figure 4 Different modes in liquid chromatography of biomolecules (A) isocratic elution, (B) linear gradient elution, (C) stepwise elution, and (D) displacement development.

The buffers used in preparative chromatography of biomolecules do not inhibit the growth of microorganisms therefore, a preservation buffer has to be applied when the column is stored. The preservation of a column is also a very delicate problem because many of the preservatives are veiy toxic for humans. Only a limited number of compounds are available for use in the production of therapeutic proteins. Sterilization of chromatography columns has been discussed... 

Although most HPLC applications utilize one or more sample preparation procedures, it is best to keep sample preparation as simple as possible [2]. Some simple procedures that have found success for protein and peptide samples are precipitation and chromatographic methods. This chapter reviews the commonly used methods and devices for sample preparation in chromatography of biomolecules, particularly proteins. The proper choice and execution of the methods can greatly affect the success of the analysis, especially in terms of recoveries [1,3,4]. Planning the sequence of procedures can minimize time and maximize efficiency a flowchart for sample preparation strategies was presented in LC-GC [5]. Table 1 summarizes some of the typical requirements for the preparation of a sample for injection onto an HPLC column. 

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