Normal phase analysis with BEH HILIC column

<p>One of my clients asked me how can she wash and care the BEH HILIC column if the column was used for normal-phase analysis with hexane and isopropanol.</p><p></p><p>For tocopherols analysis, she took the reference material as I attatched.</p><p></p><p>Thank you.</p>


  • For cleaning, Waters recommends first purging with 50:50 ACN:H2O and if this does not remove contamination, purge with 5:95 ACN:H2O.  Recommended long-term storage conditions for HILIC columns are 95:5 ACN:H2O.


  • jyang

    thank you for your reply.

    But, what I mean is for Nomal phase mode not HILIC mode as attatched file.

    In the analysis of tocopherols there was used with hexane and EA with BEH HILIC column.

    So, I wonder how can I clean and wash the column filled with hexane and EA.

    Thank you.

  • Hello,

    May I ask why you wish to clean and wash the column? Do you suspect it of being contaminated in some way?

    To answer your question regarding how to wash a normal phase (NP) column, a typical procedure would be something like:

    hexane (or whatever your NP solvent(s) is/are) > isopropanol > acetonitrile > water > acetonitrile > isopropanol > hexane (or whatever your NP solvent(s) is/are)

    The acetonitrile and water are the strong solvents in this procedure. Some people use THF during the process as well. The idea is that the solvents used that are next to each other in the process are miscible with each other.

    For storage, one normally stores a NP column in NP solvent(s) in order to minimize start-up equilibration time. Now, I must mention that we have not studied long term storage of BEH HILIC column in NP solvents. My guess is that it would be just fine but I do not have data that support that. I suggest that you store your BEH HILIC column in your preferred NP solvent(s). In the unlikely event that there is an issue with the column as a result of this storage procedure, please let me know directly and I will replace the column for you.

    I hope this information helps.


  • Hi,

    Collegues in my lab have tried to work with BEH Hilic in normal phase.

    They had a lot of trouble because retention time change over the time.

    This is due to the manufacturing process of the HILIC. Conditioning of the silica is realised with water and this is avoid in normal phase.

    We washed column with isopropanol and we also tried to dry the column in an oven but it doesn't change anything.

    At this time no acquity column is manufactured in normal phase mode.

    I know that one other manufacturer propose such a pure normal phase silica column in 1.5 µm.

    Best regards.