HPLC Separation of Amino Acids in Supplements Composition in Mixed-Mode

Amino acids are essential components of numerous formulation. Health supplements can contain various amino acids and vitamins and require quantitation of each ingredients. Amino acids are very polar compounds with limited or no retention in reversed-phase chromatography. The most common approaches are reversed-phase chromatography with ion-pairing reagent and hydrophilic interaction chromatography (HILIC). Underivatized amino acids can be retained by combination of reversed-phase and cation exchange mechanism on Primesep 100 mixed-mode. Retention time is controlled by amount of acetonitrile, buffer and buffer pH. Method does not require ion-pairing reagent. This method is for UV detection. LC/MS, ELSD or Corona CAD can be employed for analysis of amino acids with trifluoroacteic acid or ammonium formate in the mobile phase. This approach can be used for HPLC analysis of all underivatized amino acids.


Column Primesep 100, 3.2x150 mm, 5 µm, 100A
Mobile Phase MeCN/H2O - 15/85%
Buffer H3PO4
Flow Rate 1.0 ml/min
Detection UV, 200 nm


Class of Compounds Drug, Acid, Hydrophilic, Ionizable, Vitamin, Supplements, Amino acid
Analyzing Compounds Theanine, Melatonin, GABA, 5- Hydroxytryptophan

Application Analytes:
Amino Acids
gamma-Aminobutyric Acid (GABA)