Title of article :
A new on-line, in-tube pre-column derivatization technique for high performance liquid chromatographic determination of azithromycin in human serum
Author/Authors :
Bahrami، نويسنده , , Gholamreza and Mohammadi، نويسنده , , Bahareh، نويسنده ,
Issue Information :
روزنامه با شماره پیاپی سال 2006
Abstract :
Pre-column derivatization methods for high performance liquid chromatographic assay of specific pharmaceutical agents using 9-fluorenylmethyl chloroformate (FMOC-Cl) have received special attention because highly fluorescent and stable adducts are provided by these methods. However, unlike the post-column on-line techniques, long derivatization time is needed and the reaction cannot be well controlled. A new, sensitive and fast pre-column on-line derivatization technique coupled with high-performance liquid chromatography using FMOC-Cl as labeling agent is described and validated for determination of azithromycin in human serum. After extraction of the drug from serum, the residue was reconstituted in mixture of acetonitrile–phosphate buffer (3:1, v/v; pH 8.5) and directly injected onto the chromatographic system. Continuous on-line derivatization and analysis of the compounds were successfully performed using in-tube elution of FMOC-Cl. The total time needed for derivatization and chromatographic analysis of the drug was 13 min. The assay was reliable and reproducible, with limit of quantification of 10 ng/ml. The described technique may offer significant advantages over existing off-line derivatization methods using FMOC-Cl.
Keywords :
FMOC-Cl , On-line derivatization method , Reverse phase chromatography , HPLC , azithromycin , serum , Macrolide antibiotics
Journal title :
Journal of Chromatography B
Journal title :
Journal of Chromatography B