A A New Derivatization-Free Hydrophilic Interaction Liquid Chromatography (HILIC) Method for the Quantification of Phenylalanine and Tyrosine
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Abstract
In this research, a new method for determination of phenylalanine and tyrosine without derivatization based on hydrophilic interaction liquid chromatography (HILIC) was proposed. The HILIC separation was performed using guard column (3 mm x 10 mm x 3 µm) and analytical column (3 mm x 150 mm x 3 µm) with dihydroxypropyl groups bonded to silica gel as the stationary phase. The mobile phase consists of acetonitrile : 50 mM ammonium formate pH 3 at 84:16 (v/v) with a flow rate of 0.8 mL/min. Phenylalanine and a-methyl phenylalanine (internal standard) were detected at 210 nm and tyrosine was detected at 225 nm. The internal calibration curves were linear in the range of 1-500 mg/L (r2 = 0.9999). Limits of detection of phenylalanine and tyrosine were 0.7 and 0.3 mg/L, respectively. Limits of quantification of phenylalanine and tyrosine were 1 and 0.8 mg/L, respectively. Phenylalanine and tyrosine contents in eight samples of dietary supplements were analyzed using the developed method and found in the range of 149-577 and 396-499 mg/capsule with the recoveries of 95-102 and 97-102 %, respectively. The advantages of the developed method are simple, fast simultaneous analysis (within 4.5 min), and suitable for quality control in pharmaceutical and dietary supplements industry.
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