Preparative-scale chromatography flash

GENERAL CONDITIONS FOR PREPARATIVE-SCALE SEPARATIONS BY FLASH CHROMATOGRAPHY... 

When the separation procedures described in detail above are unsatisfactory for the separation of a mixture of organic compounds, purely physical methods may be employed. Thus a mixture of volatile liquids may be fractionally distilled (compare Section 2.26) or a mixture of non-volatile solids may frequently be separated by making use of the differences in solubilities in inert solvents. The progress of such separations may be monitored by application of the various chromatographic techniques detailed in Section 2.31, or indeed these techniques may be employed on the preparative scale for effecting the separation itself (e.g. flash chromatography, p. 217). The techniques of counter current distribution, fractional crystallisation or fractional sublimation (Section 2.21) may also be employed where appropriate. 

Dry-column chromatography is not a widely used today. Preparative-scale thin-layer chromatography or flash chromatography is generally preferred. Although separations are fast, the recovery of separated zones is slow and labor intensive compared with elution methods. 

Flash chromatography is widely employed for the purification of crude products obtained by synthesis at a research laboratory scale (several grams) or isolated as extracts from natural products or fermentations. The solid support is based on silica gel, and the mobile phase is usually a mixture of a hydrocarbon, such as hexane or heptane, with an organic modifier, e.g. ethyl acetate, driven by low pressure air. (Recently the comparison of flash chromatography with countercurrent chromatography (CCC), a technique particularly adapted to preparative purposes, has been studied for the separation of nonchiral compounds [90].)... 

Great skill is required to run preparative tics effectively, and even then sample obtained is often contaminated with significant quantities of sase and tic plate binding agent. Since there are now much better modem all scale separation techniques available, we do not recommend Treparative tic (see below foT details of preparative hplc and Chapter 12 for re on small scale flash chromatography). 

Within the pharmaceutical industry there has always been a need for sample purity. Any compound that is a potential drug candidate can only be fully characterised and tested once it is available in a pure form. There are many purification tools available for sample clean-up, e.g. flash chromatography, solid phase extraction, etc. 1-31. However, for the more complex purification problems where the desired compound and its associated contaminants have very similar polarities, structures, etc., preparative chromatography is the method of choice due to its superior separative capabilities. Preparative chromatography can also be scaled up from lens of milligrams to tens or even hundreds of grams of compound. The other main factor in favour of this technique is its ability to be tailored for most classes of compound. 

The use of ethyl [2-13C]acetoacetate instead of diethyl [2-t3CJmafonafe in the condensation reaction with 4H-pyran-4-one afforded ethyl 4-hydroxy[1 -13C]benzoate in 87% yield. In this case, 1.1 equiv of 4H-pyran-4-one and 1.1 equiv of potassium tert-butoxide were optimal. The addition of catalytic amounts of the base was not satisfactory. Ethyl [2-13C]acetoacetate was prepared from ethyl (2-13C]acetate as described for diethyl [2-13C]matonate.16 The maximum yield for this reaction on a 10-mmol scale was only 70% after distillation. 4H-Pyran-4-one reacted with nitromethane and potassium tert-butoxide (each 1.1 equiv) to afford 4-nitrophenol in 75% yield after purification by flash chromatography. This gives easy access to 4-nitro[4-13C]phenol. With 2,4-pentanedione, the condensation with 4H-pyran-4-one under the same reaction conditions gave 4-hydroxyacetophenone in 45-50% yield after purification. 

Many of the qualitative uses of open column chromatography have been replaced by thin layer chromatography and HPLC, and the availability of preparative HPLC systems has further reduced the use of the technique. It does, however, find continued application for the large scale separation(s) (>10g) of reaction mixtures encountered in synthetic organic chemistry, especially as with minor modifications to the basic apparatus, extremely inexpensive systems (c/. HPLC) with moderate resolution (/if >0.10) can be set up [1]. These techniques known as flash chromatography and short path chromatography are discussed in more detail herein. Gel and affinity chromatography are also still practised extensively in open column mode in the biosciences. 

Eisner, A. and Lange, R. 1993. Gram-scale preparation of plant phospholipids by flash chromatography Feff Wissenschaft Technologie-Fat Science Technology, 95, (1), 31-34. 

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