Simultaneous Determination of Free Amino Acids in Wax Apple by Reversed Phase High Performance Liquid Chromatography with Phenyl Isothiocyanate Precolumn Derivatization

A high performance liquid chromatography method for simultaneous determination of 15 free amino acids in wax apple fruit by pre-column derivatization with phenyl isothiocyanate(PITC)was established. The separation was achieved using a Atlantis T3 column under the gradient elution with the mobile phases composed of 0.1 mol/L sodium acetate buffer(pH=6.5)and acetonitrile-water(4:1)at the flow rate of 1.0 mL/min,column temperature of 35 ℃ and UV detector wavelength at 254 nm.The calibration curves of amino acid component had good linearity over the range from 2.0 to 500 mg/L with correlation coefficient all above 0.99. The detection limits were in the range of 0.08~0.35 mg/L and the quantitation limit were in the range of 0.35~1.25 mg/L. The spiked recoveries were between 85.61% and 106.68% and the RSDs were less than 5%. This method could separate the 15 amino acids with high accuracy,high sensitivity and recovery. The method was successfully used to analyze the free amino acids in wax apple fruit.