Peaks eluting in void volume region.

Chromatography Forum: LC Archives: Peaks eluting in void volume region.
Top of pagePrevious messageNext messageBottom of pageLink to this message  By ac on Saturday, December 4, 1999 - 10:27 am:

Can anybody tell how to separate the peaks eluting in the void volume region in RP-HPLC?


Top of pagePrevious messageNext messageBottom of pageLink to this message  By Anonymous on Saturday, December 4, 1999 - 01:55 pm:

Use less organic.


Top of pagePrevious messageNext messageBottom of pageLink to this message  By ac on Tuesday, December 7, 1999 - 06:59 am:

Any other suggestions.


Top of pagePrevious messageNext messageBottom of pageLink to this message  By Anonymous on Tuesday, December 7, 1999 - 10:37 am:

100% water can be used with a column that can handle it (i.e. Aquacil), slow down the flow rate, derivitize the analyte, consider using normal phase?


Top of pagePrevious messageNext messageBottom of pageLink to this message  By Anonymous on Tuesday, December 7, 1999 - 01:59 pm:

What is the composition and pH of your mobile phase?


Top of pagePrevious messageNext messageBottom of pageLink to this message  By B.Buglio on Tuesday, December 7, 1999 - 06:42 pm:

You need to try normal phase chromatography- a
silica gel column and hexane/ipa mobile phase to
start off with.


Top of pagePrevious messageNext messageBottom of pageLink to this message  By Anonymous on Wednesday, December 8, 1999 - 02:01 pm:

Without knowing your sample and current operating conditions it is difficult to advise you, however here are some general guidelines to follow assuming you are working under reversed phase conditions and your sample is not retained due to extreme polarity:
1.) Decrease your solvent strength, try methanol instead of acetonitrile or decrease your organic (maintain at least 5% with a conventional C18 column).
2.) Try a more retentive stationary phase:
a.) If you are currently using a C8, try a C18.
b.) If you are already using a C18, try one with a higher carbon loading such as Betasil C18 or BetaMax Neutral (Keystone Scientific, Inc.)
3.) Try a stationary phase that is specifically engineered for retention of small polar molecules such as AQUASIL C18, PRISM RP, and BetaMax Acid (all from Keystone Scientific, Inc.) They bonded phases have polarity incorparted into the bonded phase for added retention of polar molecules.
4.) Try a different bonded phase such as phenyl or cyano.
5.) If your sample is acidic or basic, adjust the pH so that the analyte is neutral (low pH for acids and high pH for bases). Note that inorder to operate at pH, specialty non-silica based columns are recomended.
6.) Try normal phase or another mechanism such as ion exchange.


Top of pagePrevious messageNext messageBottom of pageLink to this message  By Anonymous on Thursday, December 9, 1999 - 07:17 am:

Changing flow rate will not affect the relative retention of your analyte. The retnetion tiem fo the solvent front and analyte will be proportional to the flow rate, but capacity factor and selectivity will not be affected.


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