Don’t be a Drip: Perform Chromatography in a Flash

In this blog, I discuss the differences between open column and flash chromatography. The process of purifying compounds involves many steps; therefore, speeding up any part of the process greatly increases efficiency.

This week, I finally had fiber optic high-speed broadband installed at my house, and the difference is huge. I didn’t appreciate the improvement it would make, not to mention the joy of my nieces and nephews, who are avid gamers. They no longer complain about the lag that resulted from the high ping rate of my old setup. It reminded me of the first time I was introduced to flash chromatography. Having previously only performed gravity-fed open-column chromatography at university, I was unaware of the benefits flash chromatography introduced. I witnessed a similar reaction recently when we introduced Padma to the latest chromatography equipment at BUCHI.

Having previously only performed gravity-fed open-column chromatography at university, I was unaware of the benefits flash chromatography introduced.

I was somewhat surprised to hear that Padma was introduced to chromatography in the same manner as me many years ago. In fact, it is a testament to the impressive work achieved by Mikail Tsvet, who introduced the process at the beginning of the 20th century – a topic I discussed in a previous blog I wrote. Open-column chromatography remains the cheapest way to purify compounds, which accounts for it still being used in academic institutions that cannot afford flagship instruments that carry heavy price tags. Although the process is still a great way of introducing people to chromatography, comparing the method to modern processes is much like comparing dial-up internet with modern-day broadband. Modern setups are not only faster but also far more efficient.

To better understand the improvements flash chromatography made to the process of purifying compounds, it is important to understand the steps required to achieve a successful and efficient separation. The first step is to synthesize the molecule or to extract the molecule from a matrix.

Why would you synthesize a molecule?
Synthesis is the process of creating a desired chemical compound from simpler starting materials through chemical reactions. This is used when you need to obtain a specific compound that is not readily available in nature or when you want to produce it in a pure form. This step involves designing a series of chemical reactions that transform the starting materials into the target compound. This approach is often used in research, pharmaceuticals, and chemical manufacturing.

How do you extract a molecule from a matrix?
Extraction is the process of selectively isolating a compound of interest from a complex mixture, such as a natural source (e.g., plant material), a biological sample (e.g., blood), or a chemical mixture. The process of extraction relies on differences in the solubility of the compound of interest in different solvents. By carefully choosing appropriate solvents and conditions, you can selectively dissolve the compound you want while leaving impurities behind. For example, if you’re extracting a natural product from plant material, you might use a solvent like ethanol to dissolve the compound of interest and leave behind the plant’s cellulose and other impurities. This approach is common in natural product chemistry, environmental analysis, and forensic science, among other fields.

After one of these approaches, further purification steps are required to achieve the desired level of purity. Generally, the next step is evaporation to get rid of the solvents required for synthesis/extraction. The best way to achieve this is by using a rotary evaporator , as it performs the evaporation under a vacuum at a reduced boiling point, which prevents the molecules of interest from becoming damaged.

After evaporation, often analytical chromatography is performed whereby the crude mixture is analyzed by Thin-Layer Chromatography (TLC) or High-Performance Liquid Chromatography (HPLC). This step gives a good indication as to which method to apply for the subsequent steps. The next step is to perform preparative chromatography to further purify the compound of interest, and once again, this step is followed by an evaporation step, either through rotary evaporation or freeze-drying. Finally, the molecule needs to be analyzed to prove the presence of the molecule or confirm its purity.

How can you speed up the chromatography process?
As you can see, that’s a whole lot of steps, and to achieve the kind of purity required for specific pharmaceuticals, this process has to be repeated multiple times! As Padma and I can tell you, if you’ve ever performed open-column chromatography, you understand how slow the process is. You must first pack and prepare the column, introduce your solvent, and then wait as gravity slowly drips the solvent through the stationary phase. Drip………Drip…………Drip…….. Now imagine I’d written the word drip a few thousand more times, and you get an idea for the process. It didn’t take long for chemists to realize that this process needs speeding up. So, chemists did, as chemists do -they attached a great big pressurized air system to the column to blast the liquid through. This did work and managed to speed up the process; however, pressurized air, glass columns, and volatile solvents were obviously a recipe for disaster. It was the patent for this initial setup that introduced the term ‘flash chromatography’ –named due to the increased speed of the process, not the likelihood of explosion!

What are the benefits of flash chromatography?
Again, improvements were made, which brings us to where we are today with high-power pumps that can be accurately set to optimize the process according to the phases and compound of interest. Obviously, using a pressurized system gets things moving quicker, but there are other advantages to flash chromatography. There is no longer the need to manually pack columns – a time-consuming process that poses a health hazard due to the loose silica. Pre-packed flash columns can be used that have also been optimized for numerous workflows and compounds of interest. Modern instruments can also be heavily automated, which further improves efficiency and reproducibility. These instruments are compatible with a wide range of consumables and detection options that are often integrated into the system.

From an environmental standpoint, modern flash chromatography systems also help as they use less solvent. Less solvent means there is less to evaporate, saving even more time and resources. Due to the higher pressures, smaller particle sizes can be used in flash columns, which leads to higher-resolution separations. This negates some of the need to rework the steps of the purification process as much. Seeing Padma witness just how fast and efficient flash chromatography is makes me wish that we could have been introduced to it sooner. Although open-column chromatography is great for introducing people to the concept of chromatography, it does not show people how it is actually being performed in labs across the globe. Thankfully, more budget-friendly modular options are coming to market that provide the basics of flash chromatography, such as a powerful pump, automation, and detection solutions that may save future generations from the tedium of drip drip drip chromatography.

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