Ascorbic acid (AA) functions as an antiascorbutic agent and is a cofactor in a variety of enzymatic processes. As antioxidant it is used in pharmaceutical manufacturing and in the food industry. When exposed to oxidant stress, AA is reversibly converted to dehydroascorbic acid (DHA); then DHA is degraded to 2,3-diketogulonic acid and to other inactive species. AA and DHA are difficult to quantify due to their instability and their highly polar character. In contrast to AA, DHA has a weak UV absorption. In order to increase the sensitivity for DHA, derivatization prior or after the chromatographic separation is convenient. The present communication describes the development and validation of a reversed-phase ion pair high-performance liquid chromatography method for the separation and quantitation of DHA, AA and paracetamol. The UV-absorptivity of the DHA was enhanced by precolumn derivatization with 4,5-dimethyl-1,2-phenylenediamine. The derivatization reaction was carried out under mild conditions (10 min at ambient temperature in the dark) and the resulting DHA-quinoxaline derivative was monitored at 360 nm. The chromatographic separations were performed on a Phenomenex Synergi 4u Hydro-RP (150 x 4.6 mm) under isocratic elution conditions, using hexadecyltrimethylammonium bromide as ion-pairing reagent in the mobile phase. The validated method was successfully tested on pharmaceuticals.

Development and validation of LC method for the determination of ascorbic acid and dehydroascorbic acid in pharmaceuticals

GIOIA, MARIA GRAZIA;GATTI, RITA
2007

Abstract

Ascorbic acid (AA) functions as an antiascorbutic agent and is a cofactor in a variety of enzymatic processes. As antioxidant it is used in pharmaceutical manufacturing and in the food industry. When exposed to oxidant stress, AA is reversibly converted to dehydroascorbic acid (DHA); then DHA is degraded to 2,3-diketogulonic acid and to other inactive species. AA and DHA are difficult to quantify due to their instability and their highly polar character. In contrast to AA, DHA has a weak UV absorption. In order to increase the sensitivity for DHA, derivatization prior or after the chromatographic separation is convenient. The present communication describes the development and validation of a reversed-phase ion pair high-performance liquid chromatography method for the separation and quantitation of DHA, AA and paracetamol. The UV-absorptivity of the DHA was enhanced by precolumn derivatization with 4,5-dimethyl-1,2-phenylenediamine. The derivatization reaction was carried out under mild conditions (10 min at ambient temperature in the dark) and the resulting DHA-quinoxaline derivative was monitored at 360 nm. The chromatographic separations were performed on a Phenomenex Synergi 4u Hydro-RP (150 x 4.6 mm) under isocratic elution conditions, using hexadecyltrimethylammonium bromide as ion-pairing reagent in the mobile phase. The validated method was successfully tested on pharmaceuticals.
Abstract book and Final Programme
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119
M.G.Gioia; P.Andreatta; S.Boschetti; R.Gatti;
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11585/46992
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