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May/June 2011
order of 10kg/day) which can be used in a range of applications from reclamation of high value materials from recrystallisation liquors for example to applications involving continuous processing activities. Here the cost advantages of not having an expensive stationary phase are very attractive. Add to this the ability to process materials containing particulates, the retention time reliability that arises because of the predictability of liquid phase partition and the possibilities to run semi-continuously and we have a very exciting capability.
A photograph of a prototype large scale HPCCC instrument is presented in Figure 5 to allow the reader to get an appreciation of the size of the equipment which is capable of producing material at a rate of approximately 10 kg per day.
Alternate operating strategies An interesting enhancement to CCC already mentioned in the introductory overview is the use of intermittent counter-current extraction (ICcE) mode which takes advantage of having two liquid phases – either of which can be used as the mobile phase [11]
. With ICcE the Figure 4 A: Separation of a target peak from a crude mixture - CCC separation Figure 4 B: Separation of a target peak from a crude mixture -HPLC analytical separation of input materials Figure 4 C: Separation of a target peak from a crude mixture - HPLC analytical separation of pooled fractions isolated by CCC
techniques at the lab scale where CCC acts to enhance overall preparative capability. The objective here is to generate a first pass approach for preparative chromatography that will provide a separation solution in the fastest timescale possible with near 100% success rate.
The aim is to maximise laboratory operating efficiency by removing the requirement to ‘hand craft’ that small proportion of separations that will not yield to existing approaches.
The second is to provide a large scale, cost competitive, preparative capability (of the
two phases (mobile/stationary) are continuously alternated with the sample being continuously injected into the middle of the column or between columns if a standard 2 bobbin CCC instrument is used – this allows either the separation of binary mixtures or the concentration of a selected compound from a complex mixture while impurities are washed away. Figure 6 illustrates this process diagrammatically. Sample is continuously loaded between the two columns and flow switched regularly between reverse and normal phase modes. Under optimised conditions the target peak is held inside the instrument and gradually increases in concentration while the impurities are washed away in either the upper or lower phase. This process is illustrated by the column fraction photos which have been obtained for the preparative isolation of the target in the complex mixture application discussed earlier. These photos nicely illustrate the fractionation of this very crude material. This is confirmed by the analytical HPLC (Figure 7). With ICcE, column loading and yield is substantially enhanced compared to isocratic elution while solvent consumption is reduced making it the mechanism of choice for large scale and continuous operations.
This alternative elution method is expected to offer greatly enhanced loading and substantial reduction in solvent use compared to conventional CCC and is the subject of intensive investigation by
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