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8 August / September 2016


The next example (Figure 5) is the chromatographic separation of Efavirenz antiretroviral and its impurities on the GreenSep™ Basic. GreenSep™ Basic is inherently deactivated (10) and as such no mobile phase additive was required even though Efavirenz impurity #5 is an aromatic amine. The mobile phase for this separation was just carbon dioxide and methanol.


Figure 4: Separation of Caffeine Analogue Mixture on GreenSep™ Silica


The final application example is the chromatographic separation quinine mixture related compounds on the GreenSep™ Naphthyl (Figure 6). The quinine mixture is essentially a diastereomeric mixture and shows the separation capabilities of a naphthalene bonded phase. However, this phase is not inherently deactivated like the GreenSep™ Basic or GreenSep™ Ethyl Pyridine. The quinine and its related compounds contain amino groups as well as a base heterocyclic aromatic quinoline structure making them rather basic and so an additive (DEA) was needed improve the peak shape in this case. It should be noted, that the separation of diastereomers can be extremely challenging and in some cases will require the use of additives. In this regard we are investigating inherently deactivated stationary phases for the improved separation of diastereomers without additives. If such a stationary phase is developed it will replace GreenSep™ Naphthyl in the screening kit.


Conclusion Figure 5: Separation of Efavirenz antiretroviral and its impurities on GreenSep™ Ethyl Pyridine Figure 6: Quinine mixture related compounds on the GreenSep™ Naphthyl.


A six column screening kit for SFC has been developed (Table 2), with particular attention being paid to the preparative SFC chromatographer. The columns for the screening kit were selected based on three published studies and the important points from all three studies used to select the columns for the screening kit. All the stationary phases selected for the kit are scalable to larger column formats and different particle sizes. In addition, the selected columns are manufactured using robust support materials, refined chemical bonding procedures, are stable bonded phases and utilise high performance column packing technology. It is important not to treat the column screening kit as ‘fixed’; it should be adapted and changed as new stationary phases are developed, new separation challenges occur such as the separation of fluorine-containing pharmaceutical entities (22) or new chemometric approaches are introduced (23). However, it is important that any new columns for the screening kit must be scalable to larger column formats and different particle sizes and meet the robustness requirements of the current screening kit.


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