Controlling the retention and separation of water–soluble vitamins on zwitterion column by hydrophilic interaction echanism

Chromatographic conditions including pH, type and concentration of buffer for separation of five B vitamins on ZIC-HILIC column were investigated. Ammonium formate, ammonium acetate and ethylendiamonium chloride were used as buffer salts in the study. The results revealed that pH and buffer identity had significant effects on charged analytes. Indeed, ethylenediamonium competed very well with positively-charged analytes when the excessive retention was the problem while pH could control the charge state of the vitamins. The quanlity of acetonitrile was also employed to control the elution power in general. The mobile phase containing 10 % 40 mM ethylendiamonium chloride at pH 2.5 and 90 % ACN by volume was found to be the most suitable to separate the vitamins. The calibration curves had the R2 > 0.997, % RSD of retention time and peak areas in most of the cases were lower than 1% and 3.2 %, respectively. The developed method was applied to determine the B vitamins in Trivita BF, a commercial pharmaceutical product.