v called as “medium pressure chromatography” “An air pressure driven hybrid of medium and short column chromatography optimized for rapid separation" Popularized by Clark Still of Columbia University An alternative to slow and often inefficient gravity-fed chromatography

1. Flash chromatography
Prepared by :
SHINDE GANESH SHASHIKANT
PRAVARA RURAL COLLEGE OF
PHARMACY,PRAVARANAGAR

2. FLASH CHROMATOGRAPHY
 Also called as “medium pressure chromatography”
 “An air pressure driven hybrid of medium and short
column chromatography optimized for rapid separation"
 Popularized by Clark Still of Columbia University
 An alternative to slow and often inefficient gravity-fed
chromatography

3. Introduction..
 Differs from the conventional technique in 2 ways:
 Slightly smaller silica gel particles (250-400 mesh)
are used, and
 Due to restricted flow of solvent caused by the small
gel particles, pressurized gas (10-15 psi) 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.

4. Column vs Flash Chromatography
Traditional Column
chromatography
• Glass columns with silica
gel
• Separation is very slow
(typically many hours)
• End of the run, silica gel
must be removed,
cleaned, dried and re-
packed
• Both time consuming
and hazardous
Flash chromatography
• Pre-packed plastic
cartridges
• Solvent is pumped
through the cartridge
• Much quicker and more
reproducible
• Remaining solvent
flushed out of the column
using pressurized gas

5. Column chromatography Flash chromatography

8. Modern flash chromatography systems are sold as pre-packed
plastic cartridges, and the solvent is pumped through the
cartridge.
Systems may also be linked with detectors and fraction
collectors providing automation.
The introduction of gradient pumps resulted in quicker
separations and less solvent usage.
Flash chromatography is not expected to provide the resolution
or reproducibility of HPLC; it is a technique that can quickly
improve the purity of samples to an acceptable level.

9. SELECTION OF STATIONARY PHASE
• Smaller particles (higher mesh values {70-230} ) are used for flash
chromatography; larger particles (lower mesh values {230-400}) are
used for gravity chromatography.
• The amount of silica gel depends on the Rf difference of the
compounds to be separated, and on the amount of sample.

10. Adsorbents
• Silica: Slightly acidic medium. Best for ordinary compounds, good
separation is achieved.
• Florosil- acidic/neutral
• 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 non-polars lowest.

11. Quantity of silica gel required..
 40-63 µm silica gel particles are used
 Amount depends on 2 factors :
– Rf difference of the compounds to be separated
– Amount of sample
 ↑ silica gel -↑ the length of time for chromatography
 For,
– Easier separations, ratios closer to 30 : 1 are effective
– Difficult separations, more silica gel is often required

12. Selection of solvent system
Solvent system
Compound should have TLC Rf of 0.15 to 0.20 in the solvent
system
 Binary solvent system –
 Polarity can be adjusted
 Rate of elution can be determined
Common solvents used :
dichloromethane/hexane, ether/hexane, hexane/ethyl
acetate, and dichloromethane/methanol
High polarity of solvents increase the rate of elution of all
compounds.

13. Solvent Systems
• Flash column chromatography is usually carried out with a mixture
of two solvents, with a polar and a nonpolar component.
• 1.Hydrocarbons: pentane, petroleum ether , hexanes.
• 2. Ether and dichloromethane (very similar polarity)
• 3. Ethyl acetate

14. Two-component Solvent System
• Ether/Petroleum Ether
• Ether/Hexane
• Ether/Pentane
• Ethyl Acetate/Hexane
• Methanol/Dichloromethane
.

15. Columns
 Glass Columns :-
• A wide range of columns offer maximum flexibility for every
situation.
• Depending on the nature and the quantity of the sample offers a
series of column types which vary in form, size and performance.
.

16. COLUMN
 Plastic +Glass Column :-
• Plastic+Glass-coated Glass Columns are available for larger amounts
of samples and higher pressure applications on a high safety level.
.

17. Packing the column
 Glass column / plastic cartridges
 Has either a glass frit or a plug of cotton wool directly above the
stopcock (To prevent the silica gel from escaping from the
column through the stopcock)
 ~1/2 inches layer of clean sand above the plug of glass wool
 Make sure that surface is flat
 Pour in the silica gel using a funnel.(Do this step in hood)

18. Method
• A solvent is chosen which gives good separation and moves the
desired component to Rf = 0.20 on analytical TLC
• A column of the appropriate diameter is selected and filled with 5-6
in. of dry 40-63 µ silica gel
• Column is solvated
– The column is filled with solvent and pressure is used to rapidly
push all the air from the silica gel
• The sample is applied and the column is refilled with solvent and
eluted at a flow rate of 2 in./min
• Top of the column should never run dry

19. Procedure for Microscale Flash Column
Chromatography
 In microscale flash chromatography, the column need neither a
pinchclamp or a stopcock at the bottom of the column to control
the flow, nor does it need air-pressure connections at the top of
the column.
 Instead, the solvent flows very slowly through the column by
gravity until you apply air pressure at the top of the column with
an ordinary Pasteur pipet bulb.

20. (1) Prepare the column.
1. Plug a Pasteur pipet with a small
amount of cotton; use a wood applicator
stick to tamp it down lightly.
2. Take care that you do not use either
too much cotton or pack it too tightly. You
just need enough to prevent the
adsorbent from leaking out.
Add dry silica gel adsorbent, 230-400
mesh -- usually the jar is labeled "for
flash chromatography." One way to fill
the column is to invert it into the jar of
silica gel and scoop it out . . .

21. . . . then tamp it down before scooping
more out
Another way to fill the column is to pour the
gel into the column using a 10 mL beaker.

22. (2) Pre-elute the column.

23. (3) Load the sample onto the silica gel column
Two different methods are used to load the column: the wet
method and the dry method:
Wet loading method
The sample to be purified (or separated into components) is dissolved in a small amount
of solvent, such as hexanes, acetone, or other solvent. This solution is loaded onto the
column.

24. Dry loading method

25. (4) Elute the column.

26. 5) Elute the column with the second elution solvent.
If separating a mixture of one or more compounds, at this point
change the eluting solvent to a more polar system, Elution would
proceed as in step (4).
(6) Analyze the fractions.
If the fractions are colored, simply combine like-colored fractions,
although TLC before combination is usually advisable.
If the fractions are not colored, they are analyzed by TLC
(usually). Once the composition of each fraction is known, the
fractions containing the desired compound(s) are combined.

28. Instrumentation of Flash Chromatography

29. Advantages
• Large quantities of the sample can be separated (0.5-2g)
• Fast ( 1o to 15 minutes)
• Cost efficient
• Elaborate equipment and the purchase of expensive
equipment is not necessary
• If high resolution is required, flash chromatography is
carried out before HPLC to avoid contamination of the
expensive plates

30. Applications
• Purification of various peptides, antibiotics
• Separation of closely related organic compounds
• Purification of closely related drug intermediates
• High speed fractionation of natural products – tocopherols,
alkaloids, lignans, xanthones, stilbenes, flavonoids
• Drug discovery
• Agrochemistry
• Petrochemistry

31. Conclusion