Analysis of Amino Acids by Pre-Column Derivatization with OPA-UV Detection

June 30, 2020


Since most amino acids do not absorb or fluoresce, they need to be derivatized for high sensitivity and selectivity measurement using HPLC. Pre-column derivatization and Post-column derivatization are well-known as general methods for amino acid HPLC analysis. The pre-column method derivatizes the amino acids prior to separation on the C18 column. In the post-column method the samples are derivatized by a reagent after the separation using an ion-exchange column. JASCO can provide a dedicated HPLC system with o-phthalaldehyde (absorption detection) or dabsyl chloride (fluorescence detection) for pre-column derivatization and can also provide a system with OPA (fluorescence detection) or ninhydrin (absorption detection) for post-column derivatization.

The pre-column derivatization method has many benefits such as a simplified system configuration, a wide variety of derivatization reagents, and a high sensitivity measurement. In addition, amino acids which are derivatized using OPA can be detected by UV detector. This application note shows the result of an amino acid analysis (18 compounds) using OPA pre-column derivatization. This experiment uses the auto pre-column derivatization function of autosampler (LC-4500 model).



HPLC, UV detection, OPA pre-column derivatization, Amino acid, C18, LC-4500 Model


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Chromatography Group