Crystallisation practical techniques

Practical techniques for crystallisation have been reviewed [25,31,45,46]. Rapid screening of possible conditions are described by Carter and Carter [47] and Rayment [48], and McPherson [31] has documented nearly 200 crystallisation conditions for different proteins, providing a useful compendium which illustrates the diversity of methods employed. 

Crystallisation. The ultimate in purification of proteins or nucleic acids is crystallisation. This involves very specialised procedures and techniques and is best left to the experts in the field of X-ray crystallography who provide a complete picture of the structure of these large molecules. [A. Ducruix and R. Gieg6 eds. Crystallisation of Nucleic Acids and Proteins A Practical Approach, 2nd Edition, 2000,... 

Filtration removes particulate impurities rapidly from liquids and is also used to collect insoluble or crystalline solids which separate or crystallise from solution. The usual technique is to pass the solution, cold or hot, through a fluted filter paper in a conical glass funnel (see Vogel s Textbook of Practical Organic Chemistry, p 46). 

The above disadvantage of the lack of spatial information can be overcome by a combination of NMR data and other techniques. From the aH pulse NMR, the fraction and the molecular mobility of different molecular environment can be obtained as free induction decays (FIDs) within a short time, which is suitable for a practical, better understanding of the morphology-property relationship. Wide angle X-ray diffraction (WAXD) and small angle X-ray diffraction (SAXD) as well as electron microscopy provide direct information between the nano- and micrometre scale. A combination of NMR data with those from X-ray diffraction and electron microscopy should be able to analyse the structure from the atomic level to the macro scale. In this review, the morphology-property relationship, the dynamics of morphological transition, the kinetics of crystallisation, etc. analysed by a combination of NMR and other tools are introduced. 

Differential Scanning Calorimetry (DSC) and Thermogravimetric Analysis (TGA) of amoxicillin trihydrate showed loss of the water of crystallisation from about 50 to 150 °C followed by decomposition of the amoxicillin from about 160°C upwards [18], DSC and TGA of amoxicillin sodium salt in an oxygen atmosphere showed loss of water at about 100°C and thermal decomposition in multiple events from about 200 to 500°C [19]. The techniques are of little practical value for these compounds. 

Main Section 6.2 was compiled as a theoretical introduction to the more practical problems of obtaining X-ray crystal structures of biological macromolecules. In fact, although X-ray diffraction has proved immensely popular as a means of biological macromolecule structure determination, the technique is still quite onerous to implement and the possibility for error surprisingly high. These problems will become clear in this section. The first and still the most difficult problem is that of biological macromolecule crystallisation. If there are no well formed crystals then there can be no X-ray crystal diffraction ... 

A simple method of preparing single crystals is to melt the material and to crystallise it by slow cooling but practically all the ferrites dissociate at temperatures below their melting points, with loss of O2. To overcome this problem several techniques have been used which are based on the formation of a flux, a solution, or a volatile chemical compound to obtain ferrite single crystals. 

Crystallisation. The ultimate in purification of proteins or nucleic acids is crystallisation. This involves very specialised procedures and techniques and is best left to the experts in the field of X-ray crystallography who can provide a complete picture of the structure of these large molecules. [A. Ducruix and R. Giege Eds, Crystallisation of Nucleic Acids and Proteins A Practical Approach, 2nd Edition, 2000, Oxford University Press, ISBN 0199636788 (paperback) T.L. Blundell and L.N. Johnson Protein Crystallisation, Academic Press, NY, 197, A. McPherson Preparation and Analysis of Protein Crystals, J.Wiley Sons, NY, 1982, A. McPherson, Crystallisation of Biological Macromolecules, Cold Spring Harbour Laboratory Press, 2001 ISBN 0879696176, see also Bibliography in Chapter 1.]... 

So far, attempts at putting this attractive idea into practice have not been very succesful (56). This is due to the rate of segregation of the phases, which proved to be too slow. However, the method is so promising that it is to be hoped that techniques for overcoming this drawback will soon be invented. As matters stand now. this is the only approach by which non-crystallisable polymers can be separated into fractions with such narrow distributions and, at the same time, is suitable for application under truly preparative conditions. 

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