4,7-Phenanthroline-5,6-dione (phanquinone) was used as a fluorogenic labelling reagent in pre-column derivatization for the quality control of amino acids in pharmaceuticals. The amino acids adducts were efficiently separated by C12 RP HPLC using a ternary mixture of triethylamine (TEA) phosphate buffer (pH 2.5, 0.05M)-methanol-THF as mobile phase by varying composition gradient elution and detected fluorometrically. The results obtained by the proposed method were compared statistically, by means of the Student’s t-test and the variance ratio F-test, with those obtained by a rapid reference method, which involved o-phthaldialdehyde (OPA) as pre-column reagent; no significant difference was found. The stronger derivatization conditions (60°C, pH 8, 60 min) required for the method with phanquinone are compensated by the major stability of derivatives and by the absence of fluorescent degradation products.
HPLC-Fluorescence determination of amino acids in pharmaceuticals after pre-column derivatization with phanquinone
GATTI, RITA;GIOIA, MARIA GRAZIA;
2004
Abstract
4,7-Phenanthroline-5,6-dione (phanquinone) was used as a fluorogenic labelling reagent in pre-column derivatization for the quality control of amino acids in pharmaceuticals. The amino acids adducts were efficiently separated by C12 RP HPLC using a ternary mixture of triethylamine (TEA) phosphate buffer (pH 2.5, 0.05M)-methanol-THF as mobile phase by varying composition gradient elution and detected fluorometrically. The results obtained by the proposed method were compared statistically, by means of the Student’s t-test and the variance ratio F-test, with those obtained by a rapid reference method, which involved o-phthaldialdehyde (OPA) as pre-column reagent; no significant difference was found. The stronger derivatization conditions (60°C, pH 8, 60 min) required for the method with phanquinone are compensated by the major stability of derivatives and by the absence of fluorescent degradation products.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
