A 10 mg L−1 mixed standard solution containing the 13 colorants was used to study the optimum conditions of separation.
The initial gradient elution program used was as follows.
For the initial 2 min, the mobile phase A (ammonium acetate buffer) goes through the column for elution of inorganic compounds.
Between 2 and 25 min, a gradient is carried out and for the last 2 min (25–27 min) the mobile phase B passes through the column for the elution of possible organic impurities. Finally, a 2-min equilibrium phase of the column is taking place for the
next run.