The facility uses the Waters Pico-Tag System for the analysis of amino acids. It involves the hydrolysis and pre-column derivatization of the hydrolyzates using PITC followed by reverse phase HPLC. Samples are dried in pyrolyzed borosilicate tubes in a vacuum centrifugal concentrator and subjected to vapour phase hydrolysis by 6N HCl with 1% phenol at 110°C for 24 hours under pre-purified nitrogen atmosphere.
After hydrolysis, excess HCl is removed by vacuum, hydrolyzates washed with redrying solution and derivatized with phenyisothiocyanate (PITC) to produce phenylthiocarbamyl (PTC) amino acids. The derivatization method is accomplished at room temperature, derivatizes both primary and secondary amino acids (i.e. proline and hydroxyproline) and forms very stable amino acid derivatives detectable at low picomole levels.
Derivatized amino acids are redissolved in phosphate buffer and transferred to injection vials which are loaded into the autosampler for automatic injection.