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HPLC column - 2005/11/28 19:11
How you clean a column, to a great extent depends on what you are trying to clean off of it. I have seen some chemicals used that I would have been reluctant to try on my own for fear they would damage the column. One must also consider the column itself, substrate and C-18 bonding. A silica substrate without end capping versus, non-polar end capping versus the new polar end capping columns. There are also C18 columns with substrates other than silica. The simplest most strate forward cleaning that I use for most of my reverse phase columns is with a 3 solvent gradient HPLC. I use a program with the following parameters. Start at 33%H2O:33%MeOH:34%MeCN flowing at 1ml/min. Over 30 minute periods change to 100% H2O, then to 100% MeOH and finally to 100% MeCN. Then over a 15 minute period return to starting conditions. Set the instrument to run this program 3 times overnight.
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HPLC column - 2005/11/28 19:12
We often clean our column first 40 min with water, then 40 min with methanol, then THF, again methanol and next water. When you use a higher temperature this works better. Do not couple your column onto the detector, because if its really sticky material, its possible that the component blocks your detector cell, so you have to change this. Sometimes it helps when you clean and use your column in reversed way.
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HPLC column - 2005/11/28 19:13
Definitely, never clean a column feeding a detector.
Be very careful about cleaning columns with reverse flow. They are like cats. They do not mind being scratched backwards but will sometimes react violently to being stroked backwards. If you do back flush, do so with very slow flows and absolute minimum back pressure.
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Re:HPLC column - 2006/06/12 18:33
SGE HPLC fittings, guard cartridges and in-line filters are completely compatible with all other manufacturers' HPLC columns and systems.
If you are doing any method development with reversed phase HPLC, Wakosil II 5C18 RS columns should be the first columns to try.
Always use a guard cartridge to prolong column life.
For quick and easy column connections use EasyLOK nuts.
If your reversed phase HPLC run is taking too long on a C18 or ODS column try using a SGE Wakosil II 5C8 RS (basic compounds and method development), Exsil C8 (existing method on an older style ODS), or Nucleosil C8 (protein analysis and peptides).
SGE Guard cartridges can be used in conjunction with competitors' columns (for those customers with methods validated on other columns).
PEEKsil tubing is an excellent alternative to PEEK tubing as its internal bore is unaffected by any organic solvents.
Wakosil II 5C18 AR columns have a pH range of 1.4 - 9.4 making it a very TOUGH phase.
If you are close to the limit of detection using a standard 4.6mm or 4mm ID HPLC column try using a 2mm ID column. This saves solvent too.
For complete loop fill, the syringe capacity should be greater than twice the loop volume. The loop capacity sets the injection volume. For partial loop fill, the injection volume should be no greater than half the loop capacity. The injection size sets the injection volume.
If you can't pre-filter samples, make sure to use a low dead volume in-line filter after the injector.
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