Have you heard about flash chromatography? Is it just another name for high-performance liquid chromatography (HPLC)? And if not, what is the difference between the two? These are some of the very common, mind-boggling questions for anyone who is trying to learn chromatography anew. Well, these questions are genuine enough because flash chromatography is a relatively less-discussed technique even among chromatographic experts.
Flash chromatography is quite similar to high-performance liquid chromatography but slight differences still exist between the two techniques which help us in drawing a fine line between them. Thus, in this article, we will discuss everything you need to know about flash chromatography.
What is flash chromatography
Flash chromatography is a type of liquid column chromatography in which the solute components travel down the stationary phase using air pressure. It is pre-dominantly a preparative chromatographic tool rather than being an analytic one. Flash chromatography is different from other types of chromatography because the others largely operate on gravitational pull.
Flash chromatography is usually performed in plastic cartridges packed with silica gel particles of size 40-60 µm. An organic solvent is used as the mobile phase under mildly pressurized (50-200) Psi conditions provided by a compressed gas. Flash chromatography is also sometimes known as a medium-pressure chromatography.
Historical background of flash chromatography
Flash chromatography was introduced for the first time back in 1978 by Clark Still. However, it was not until 1994 that disposable plastic cartridges were introduced and flash chromatography became a viable purification tool. The technique was named flash chromatography because it facilitates component separation in a ‘flash’’ i.e., rapidly.
How to perform flash chromatography
Below is a step-by-step guide on how is flash chromatography performed in a laboratory.
Step I: Column packing
The column in flash chromatography is a plastic cartridge packed with a pure, solid medium such as small sized silica particles. These cartridges are usually bought pre-packed from the market but they can also be packed in the laboratory. The cartridge packing may vary depending on sorbent weight, particle diameter as well as on the stationary phase functionalities (normal and/or reverse-phase) as per analyte requirement.
If you want to know more on these different types of stationary phase materials, we recommend: normal-phase and reverse-phase chromatography.
Step II: Solvent pumping
The mobile phase solvent is then pumped into the column under a medium pressure provided by a compressed gas such as nitrogen or air.
Step III: Sample loading
The complex sample mixture to be separated is then introduced from the top of the column. The sample components get separated on the basis of their solubility in the mobile phase solvent. Highly soluble components get pushed down the column faster while the less soluble ones stay behind. Thus, component partitioning occurs.
Step IV: Elution
Low viscosity, organic solvents such as n-hexane, ether, dichloromethane, ethyl acetate and dimethyl phthalate etc. and/or their mixtures are commonly used as mobile phase eluents in flash chromatography.
Based on the difference in solubility and consequently on their differing rates of diffusion in these solvents, components from the sample mixture get eluted out of the column at different times. These components are thus separately collected. A detector response is generated for each component and plotted as a chromatogram.
Step V: Detection
The detectors for flash chromatography are same as that used for HPLC. The most popular detector choices for flash chromatography includes a UV-Visible detector, an evaporative light scattering detector (ELSD) and a mass spectrometric (MS) detector. For more information on how these detectors work, you may read our article: detectors for chromatography.
How flash chromatography is different from HPLC
The table given below shows how flash chromatography although quite similar to HPLC but is still different.
|High-performance liquid chromatography (HPLC)||Flash chromatography|
|High pressure applied approx. 6000 Psi||Low to medium pressure applied (50-200) Psi|
|Stainless-steel based column||Small, disposable plastic cartridge column|
|Comparatively larger silica (Si) gel particle size (300 µm)||Smaller particle size (40-60) µm|
|Important for chromatographic
separation and ‘analysis”
|Important primarily as a chromatographic
|Allows analysis in nano or micrograms||Allows bulk sample purification in a single run
(grams to industrial kilograms)
|It is an expensive technique||It is relatively less-expensive|
There is another liquid chromatographic variant, similar to HPLC which is called ultra-high performance liquid chromatography (UHPLC). While flash chromatography is a medium pressure liquid chromatography as compared to HPLC, UHPLC on another extreme runs on extraordinarily high-pressure conditions, as high as 20,000 Psi.
Where do we need flash chromatography
- Purification prior to analysis: Flash chromatography finds important applications for the purification of biological molecules such as proteins and antibiotics as well as in organic chemistry for preparative analysis of pharmaceutical compounds. The chemist applies flash chromatography to purify these compounds before they can be analyzed by other types of chromatography.
- High speed flash fractionation of natural products: A series of natural compounds including plant bioactives (alkaloids, flavonoids and cannabinoids etc.) can be concentrated and any impurities (oil contaminants) removed. This protects the analytical HPLC column to be used later on, from damage. Thus, flash chromatography is also called prep-HPLC.
- Removing reaction by-products: Unreacted starting materials and unwanted by-products can be removed from a reaction mixture prior to performing the next reaction step. In this way, flash chromatography aids in novel organic synthesis.
- Purification of isomers: Isomers, otherwise difficult to separate, can be separated, purified from their sample mixture (with > 98% purity) and identified by flash chromatography followed by TLC.
Flash chromatography is important because it speeds up conventional chromatographic purifications. It is considered a hybrid chromatographic technology with combined benefits of applying a medium pressure and using a shorter column. These conditions allow sample purification in a short time span.
Flash chromatography can separate several grams of compounds rapidly; 2-3 hours work via conventional purification methods can be equated to 15 minutes of flash chromatography. Thus, it definitely requires more appreciation and a louder applause in the world of chromatography.
Lastly, as is our tradition, here is a video tutorial for you on flash column chromatography.
1. Kasprowiak, A., F. Cazier-Dennin and P.-E. Danjou (2020). “Flash Chromatography System: A Practical Tool for Demonstrating the Influence of Column Characteristics on Chromatographic Resolution.” Journal of Chemical Education 97(4): 1145-1150.
2. Stevens, W. C. and D. C. Hill (2009). “General methods for flash chromatography using disposable columns.” Molecular Diversity 13 (2): 247.