To remove sodium hexanesulfonate from a Luna Omega Polar C18 column, flush with deionized water for 10–20 column volumes to remove buffer salts. Follow with 80:20 acetonitrile (ACN):water or methanol (MeOH):water to dissolve organic residues. Then, use 10–20 mM ammonium hydroxide in 50:50 ACN:water or MeOH:water to break ion-pair interactions. If residues persist, flush with 0.1% trifluoroacetic acid in 50:50 ACN:water or MeOH:water. Finally, flush with water and re-equilibrate with the original mobile phase. Perform regular gradient flushing and follow manufacturer guidelines for optimal column care to maintain performance and extend the column’s lifespan.

You may not be able to completely remove the sodium hexanesulfonate from the column surface. The column surface has been changed. The suggested generic flush and wash methods may reduce the amount of non-bound material, but they will not restore the original surface chemistry of the column back to "new" (it is a different column after use).

Label the column as used with "ion-pairing reagents" and dedicate it to the one specific application. The surface has been permanently changed and you do not want to use this column with any other samples as the results may not be reproducible. Do not use this column to develop new methods on either.

This is a basic fundamental that we teach in our HPLC training classes;

• "If you use ion-pairing regents with an HPLC column, then dedicate that column to the the specific analysis method". *If you use it to develop a new method, then the results may not be the same when you use a "new" column. Methods can not be transferred or reproduced so data may be invalid. We have seen many labs fail regulatory compliance for using the same column for different analysis methods (where one used IP reagents in the mobile phase and the other did not not).

HPLC columns are inexpensive consumables. You should always have fresh new columns available for use with new methods of analysis. If possible, avoid the use of ion-pairing regents, but if you do need to use, understand that you must dedicate the specific column to the analysis method. Also note the ion pairing regents often "stick" to other materials inside the HPLC's system flow path too (sometimes resulting in carryover contamination). The plastic parts are especially troublesome and replacement is often recommended at regular intervals to reduce contamination (e.g. injector valve rotary seals, needle seats, piston seals...).

Never encountered problems: rinse the column using a gradient starting at 20% ACN in water to 100% ACN in 30 minutes. Store the column in 100% ACN. Our system contained also PEEK tubing: no problem.