The bioactive components of many naturally occurring products are of great scientific interest and potentially enormous commercial value because of their pharmacological effects in humans and other species.
The desired components which are to be isolated from natural products often exist at low concentrations within very complex matrices and may comprise compounds which are susceptible to chemical degradation and whose molecular weights can vary from tens to thousands of Daltons. By conventional means, i.e. methods involving solid-liquid chromatography, the isolation and purification of these compounds is usually both time-consuming and expensive involving multiple processing steps to achieve the required purity. Significant losses of target compound(s) by chemical degradation and/or irreversible adsorption to the solid phase can occur at any step.
High performance countercurrent chromatography (HPCCC) is very effective for the fractionation of and the isolation of target compounds from crude natural product extracts. It can be used alone or in tandem with orthogonal preparative solid-liquid chromatography methodologies to dramatically enhance overall product recoveries and often product purities. Particularly in the bioassay-guided fractionation of plant derived compounds, HPCCC is an excellent isolation and purification tool.
The technique can handle from mg to kg quantities of crude natural product extract to provide compounds for further study. The polarity range of compounds which can be processed effectively is large but the technique is particularly suited to those of high polarity which are often problematic when solid-liquid chromatography methods are used. Typical compound groupings to which HPCCC has been successfully applied are phytochemicals (including essential oils, cannabinoids, glucosinolates, polyphenolics, terpenoids and alkaloids), mycotoxins, antibiotics, peptides, etc.
|Stationary Phase||Hexane-ethyl acetate-methanol-water (1:0.4:1:0.4 v/v)||Light petroleum-ethyl acetate-tetrachloromethane-methanol-water (1:1:8:6:1 v/v)||Zorbax Eclipse XD8-C18|
|Actual Column Volume (ml)||4500||300||17|
|Mobile Phase||Hexane - Ethyl Acetate||Methanol - Water||Methanol - Water|
|Scale Factor to 1L Col. Vol.||0.22||3.33||58.82|
|Run Time (min)||20||450||40|
|Purity (%)||> 99.9||> 98.5||> 99.0|
|Sample capacity per run per 1L of bed volume (g)||9.46||6.66||1.15|
|Productivity (mg/min/L of bed volume)||473||15||29.00|
Table - Comparison of Chromatography techniques for purifying the isomers of Houpu