Try washing the column for 2 hours with 70:30 water / acetonitrile and retest the column for peak shape.

Note: If you are using a method with a buffer such as 16mM ammonium acetate in the mobile phase and then stored the column in pure acetonitrile immediately afterwards, you may observe peak splitting when using your ammonium acetate method next time you use it.

It is possible that ammonium acetate has precipitated in the column since it is insoluble in pure acetonitrile, so it is always advisable that after use to wash the ammonium acetate out of the column before introducing it to a pure organic solvent to prevent this.

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