Column Selection

These challenges drove the development of Shimadzu’s innovative Nexera UFPLC purification system. Ultra-Fast Preparative Liquid Chromatography (UFPLC) automatically performs the various processes associated with preparative isolation of target compounds through a combination of preparative LC and trapping columns.

The process consists of three main stages:

• Separation of target compounds from a complex sample by preparative LC, followed by introduction into trapping columns for fractionation and concentration.
• Online washing of trapping columns to remove impurities and counter ions during purification.
• Elution of target compounds from the trapping columns using organic solvent.

Following fractionation, the trapping cartridges can undergo online purification procedures, including:

- Automatically by mass or other detector parameters

- By retention time

- Manually through the software interface

An example of a trapping cartridge containing concentrated target compounds and impurities prior to purification is shown in figure beside. Compared to conventional approaches, evaporation can be performed up to 90% faster, while the entire purification sequence remains fully automated.

During the elution stage, the desired compound is backflushed from the trapping cartridge using an organic solvent. This produces a highly concentrated, high-purity fraction in a solvent composition that is much easier and faster to evaporate. The purified sample can then be re-analysed on an analytical LC system to verify purity.

As illustrated here, the traditional workflow can take significantly longer to complete due to the multiple manual post-fractionation steps involved.

By reducing or eliminating these downstream processing requirements, UFPLC can deliver results up to 18 times faster than conventional preparative workflows.

For preparation and purification by UFPLC, the dedicated Purification Solution software is used. Purification Solution facilitates the entire process, from setting the preparative conditions to the liquid recovery of the target compound on a simple user interface. The Purification Solution operation screen is shown here.

The chromatogram of preparative separation, the trapping column into which fractions were introduced, and the elution chromatogram after purification are displayed on a single screen. It allows the user to confirm the entire process of preparative purification for the target compounds at a glance.

Let’s look at a real example where ibuprofen is purified on a standard preparative LC vs the Nexera UFPLC Table 1. The column was maintained between the two systems, however, the amount of load which can be collected is 5x greater on the UFPLC due to the trapping columns.

The standard system has the fraction collected in the mobile phase which is higher in aqueous solvent than the concentrated fraction which is eluted from the trapping column in high organic solvents. This reduced the drying time by over half. The quality of the powderised compound is also superior on the UFPLC after drying as the standard results in co-precipitation of mobile phase derived compounds.