Particularly in the bioassay-guided fractionation of plant derived compounds, HPCCC is an excellent isolation and purification tool. Such natural product compounds are often large molecules which exist at low concentrations and need to be separated from very complex crude samples. These crude samples can also vary appreciably in both the amount and type of contamination, and target compound present. By conventional means, the separation and purification of these molecules is usually both time-consuming and expensive. Often many process steps are required to achieve the purification, and there are often losses of the target compound at each stage of the process.
HPCCC, on the other hand, enables high-purity compounds to be obtained from unprocessed crude samples in one or two simple stages. HPCCC can be used on compounds of any polarity, but has special advantages for compounds of high polarity. This is because it circumvents the problems associated with solid-phase adsorbents and preserves the chemical integrity of natural mixtures being fractionated. It dramatically cuts processing time and cost, as well as offering a process that is easily scaled from mg to kilo scale without the need for any redevelopment.
HPCCC may be used for the isolation and purification of bioactive natural products in quantities sufficient for the study of their biological, pharmacological and clinical effects both in vitro and in vivo. It also finds applications in the screening of new bioactive compounds in crude extracts and for micro scale isolations where the key advantage is that the method involves no sample losses.
High performance counter current chromatography is very effective for the initial fractionation of crude plant extracts and can be used in tandem with preparative high performance liquid chromatography to dramatically improve overall product recoveries without any loss of product purity.
Table – Comparison of Chromatography techniques for purifying the isomers of Houpu
