Ayare P*, Khanvilkar V, Chalak N
Department of Quality Assurance,
Bharati Vidyapeeth’s College of Pharmacy, Navi Mumbai

Most of the sophisticated analytical techniques like NMR, MS and FT-IR need sample in the extreme pure form which is usually achieved by Preparative column chromatography but it is time consuming. From past few decades, technique called as Flash chromatography is developed which is a modification of Preparative column chromatography. This is an air pressure driven technique comprising of medium and short column chromatography, optimised for rapid separation of organic compounds. Modern flash chromatographic system consists of pre-packed plastic cartridges wherein solvent is pumped through the cartridges to achieve separation. These systems are also linked with detectors and fraction collectors which can even be automated. It is a simple, fast and economic approach to preparative liquid chromatography for purification of chemical species.This review highlights the principle involved, instrumentation, general procedure and advancement in Flash chromatography along with its applications.


PharmaTutor (ISSN: 2347 - 7881)

Volume 2, Issue 5

Received On: 28/02/2014; Accepted On: 25/03/2014; Published On: 01/05/2014

How to cite this article: P Ayare, V Khanvilkar, N Chalak; Flash Chromatography: Area & Applications; PharmaTutor; 2014; 2(5); 89-103

All chromatographic methods –with the exception of TLC- use columns for the separation process. Column chromatography has found its place in many laboratories for preparative purposes as well as for reaction control in organic synthesis. The importance of column chromatography is mainly due to following factors:
Simple packing procedure.
Low operating pressure.
Low expense for instrumentation.

Column chromatography is separated into two categories, depending on how the solvent flows down the column. If the solvent is allowed to flow down the column by gravity, or percolation, it is called Gravity column chromatography. If the solvent is forced down the column by positive air pressure, it is called Flash chromatography.[1] In traditional column chromatography a sample to be purified is placed on the top of a column containing some solid support, often silica gel. The rest of the column is then filled with a solvent (or mixture of solvents) which then runs through the solid support under the force of gravity. The various components to be separated travel through the column at different rates and then can be collected separately as they emerge from the bottom of the column. Unfortunately, the rate at which the solvent percolates through the column is slow. In flash chromatography however air pressure is used to speed up the flow of solvent, dramatically decreasing the time needed to purify the sample, therefore making the column and running the separation could take less than 10-15 minutes.[2]

Flash chromatography is basically an air pressure driven hybrid of medium pressure and shorter column chromatography which has been optimized for particularly rapid separation. Flash chromatography is a technique used to separate mixtures of molecules into their individual constituents, frequently used in the drug discovery process.[1]

Flash chromatography, also known as medium pressure chromatography, was popularized several years ago by Clark Still of Columbia University, as an alternative to slow and often inefficient gravity-fed chromatography. Flash chromatography differs from the conventional technique in two ways:
1. Slightly smaller silica gel particles (250-400 mesh) are used
2. Due to restricted flow of solvent caused by the small gel particles, pressurized gas (10-15 psi) is used to drive the solvent through the column of stationary phase.

The net result is a rapid (“over in a flash”) and high resolution chromatography.[3]

Automated flash chromatography systems include components normally found on more expensive HPLC systems such as a gradient pump, sample injection ports, a UV detector and a fraction collector to collect the eluent. Typically these automated systems separate samples from a few milligrams up to an industrial kg scale and offer much cheaper and quicker solution to doing multiple injections on prep-HPLC system.The software controlling an automated system coordinate the components, allow a user to only collect the factions that contain their target compound and help the user to find the resulting purified material within the fraction collector. The software also saves the resulting chromatograph from the process for archival and/or later recall purposes.[4]

The principle is that the eluent which is a liquid, under gas pressure (normally nitrogen or compressed air) rapidly pushed through a short glass column. The glass column is packed with an adsorbent of defined particle size withlarge inner diameter. The most used stationary phase is silica gel 40 – 63 μm, but obviously packing with other particle sizes can be used as well. Particles smaller than 25 μm should only be used with very low viscosity mobile phases, because otherwise the flow rate would be very low. Normally gel beds are about 15 cm high with working pressures of 1.5 – 2.0 bars. Originally only unmodified silica was used as the stationary phase, so that only normal phase chromatography was possible. In the meantime, however, and parallel to HPLC, reversed phase materials are used more frequently in flash chromatography.[3]

Chromatography exploits the differences in partitioning behaviour between a mobile phase and a stationary phase to separate the components in a mixture. Compounds of the mixture interact with the stationary phase based on charge, relative solubility or adsorption. The retention is a measure of the speed at which a substance moves in a chromatographic system. In a continuous development system like HPLC or GC where the compounds are eluted with the eluents, the retention is usually measured as the retention time (rt), the time between the injection and detection. In un-interrupted development system like TLC, the retention is measured as the retention factor (Rf), the run length of the compound divided by the run length of the eluent front. Rf = Distance travelled by the solvent front. [4]


The basic prerequisite for successful separations is the choice of the proper adsorbent. The most important stationary phase in column chromatography is silica. Silica gel (SiO2) and alumina (Al2O3) are two adsorbents commonly used by the organic chemist for column chromatography. These adsorbents are sold in different mesh sizes, as indicated by a number on the bottle label: “silica gel 60” or “silica gel 230-400” is a couple examples. This number refers to the mesh of the sieve used to size the silica, specifically, the number of holes in the mesh or sieve through which the crude silica particle mixture is passed in the manufacturing process. Adsorbent particle size affects how the solvent flows through the column. Smaller particles (higher mesh values) are used for flash chromatography; larger particles (lower mesh values) are used for gravity chromatography. For example, 70-230 silica gels are used for gravity columns and 230-400 mesh for flash columns. The amount of silica gel depends on the Rf difference of the compounds to be separated, and on the amount of sample. For n grams of sample, you should use 30 to 100 n grams of silica gel. For easier separations, ratios closer to 30: 1 are effective, for difficult separations, more silica gel is often required. However, by using more silica gel, the length of time required for the chromatography is extended. The density of powdered silica gel is about 0.75 g per mL.[1] [2]

These are some adsorbents which are mainly used in flash chromatography:-
· Silica: Slightly acidic medium. Best for ordinary compounds, good separation is achieved.
· Florisil: Mild, neutral medium. 200 mesh can be effective for easy separations. Less than 200 mesh best for purification by filtration. Some compounds stick on florisil, test first.
· Alumina: Basic or neutral medium. Can be effective for easy separations, and purification of amines.
· Reverse phase silica: The most polar compounds elute fastest, the most nonpolar slowest.[5]

The properties of commonly used flash solvents:-
· The compound of interest should have a TLC Rf of ≈0.15 to 0.20 in the solvent system you choose.
· Binary (two component) solvent systems with one solvent having a higher polarity than the other are usually best since they allow for easy adjustment of the average polarity of the eluent.
· The ratio of solvents determines the polarity of the solvent system, and hence the rates of elution of the compounds to be separated.
· Higher polarity of solvent increases rate of elution for all compounds.
· If your Rf is a≈0.2, you will need a volume of solvent ≈5X the volume of the dry silica gel in order to run your column.[4]

Solvent Systems:
Flash column chromatography is usually carried out with a mixture of two solvents, with a polar and a nonpolar component. Occasionally, just one solvent can be use. The only appropriate one component solvent systems (listed from the least polar to the most polar):
1. Hydrocarbons: pentane, petroleum ether, hexanes.
2. Ether and dichloromethane (very similar polarity)
3. Ethyl acetate.

The most common two-component solvent systems (listed from the least polar to the most polar):
4. Ether/Petroleum Ether, Ether/Hexane, and Ether/Pentane: Choice of hydrocarbon component depends upon availability and requirements for boiling range. Pentane is expensive and low-boiling, petroleum ether can be low-boiling, and hexane is readily available.
5. Ethyl Acetate/Hexane: The standard, good for ordinary compounds and best for difficult separations.
6. Methanol/Dichloromethane: For polar compounds.
7. 10 % Ammonia in Methanol Solution/Dichloromethane: Sometimes moves stubborn amines off the baseline.
8.For basic (i.e. nitrogen containing) compounds, it is sometimes useful or necessary to add a small amount of triethylamine or pyridine to the solvent mixture (about 0.1%).
9.For acidic compounds, a small amount of aceticacid is sometimes useful. In these cases, the acetic acid can often be safely rotavaped away by adding  portions of toluene and concentrating to a few mL volumes and repeating this several times. As acetic acid boils at a lower bp than toluene, this will remove the acid without exposing the neat compound to it.[1] [5]


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