Determination of gentamicin in bovine milk using liquid chromatography with post-column derivatization and fluorescence detection.
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.
Kijak
P J
PJ
Food and Drug Administration, Center for Veterinary Medicine, Beltsville, MD 20705, USA.
Jackson
J
J
Shaikh
B
B
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000032:analysis*; D002853:Chromatography, Liquid; D005839:Gentamicins / Q000032:analysis*; D007202:Indicators and Reagents; D008892:Milk / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D009764:o-Phthalaldehyde
D000903:Antibiotics, Antineoplastic; D005839:Gentamicins; D007202:Indicators and Reagents; D009764:o-Phthalaldehyde