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TITLE:  Determination of gentamicin in bovine milk using liquid chromatography with post-column derivatization and fluorescence detection
AUTHORS:  Kijak PJ;Jackson J;Shaikh B;
YEAR:  1997
JOURNAL ABBREV:  J Chromatogr B Biomed Sci Appl
JOURNAL FULL:  Journal of chromatography B, Biomedical sciences and applications
VOLUME:  691
END PAGE:  382
KEYWORDS:  analysis;Animals;Antibiotics,Antineoplastic;Centrifugation;chemistry;Chromatography;Chromatography,Liquid;Fluorescence;Food;Gentamicins;Indicators and Reagents;Methanol;Milk;o-Phthalaldehyde;Solid Phase Extraction;Spectrometry,Fluorescence;veterinary;Veterinary Medicine;Water;
ABSTRACT:  A method capable of separating and quantifying the three major and one minor components of gentamicin in milk has been developed. The method is capable of detecting 15 ng/ml gentamicin, based on a total of the four components. Milk samples are centrifuged at 4 degrees C, the fat layer removed, and the samples deproteinated with 30% trichloracetic acid. After a second centrifugation, the supernatant is passed through a C18 solid-phase extraction column. The column is washed with water, water-methanol (50:50) and methanol. Ammonium hydroxide (16%) in methanol is used to elute the gentamicin. The eluent is evaporated to near dryness and taken up with water. An aliquot of the sample is then mixed with an ion-pairing reagent for chromatography. Separation is achieved using pentanesulfonic acid in a water-methanol mobile phase on a C18 reversed-phase column. The o-phthalaldehyde fluorescence derivatives of gentamicin are formed post-column and are detected with excitation at 340 nm and emission at 430 nm. The percent recovery of gentamicin averaged 72, 78 and 88% from milk samples fortified at 15, 30 and 60 ng/ml, respectively
AFFILIATIONS:  Food and Drug Administration, Center for Veterinary Medicine, Beltsville, MD 20705, USA