Determination of clavulanic acid in calf plasma by liquid chromatography tandem mass spectrometry.
A method for the quantification of clavulanic acid in calf plasma using high-performance liquid chromatography combined with electrospray ionization (ESI) mass spectrometry, operating in the negative ionization mode (LC-MS/MS), is presented. Sample preparation includes a simple and fast deproteinization with acetonitrile and a back-extraction of the acetonitrile with dichloromethane. Chromatography is performed on a reversed-phase PLRP-S polymeric column using 0.05% formic acid in water and acetonitrile. The limit of quantification is 25 ng/ml, which is lower than other published methods using ultraviolet (UV), fluorimetric or mass spectrometric detection. The limit of detection is calculated to be 3.5 ng/ml. The stability of clavulanic acid was demonstrated according to The Guidelines of Bioanalytical Method Validation of The Food and Drug Administration (FDA): freeze and thaw stability, short-term stability, long-term stability, stock solution stability and postpreparative stability. The method is used in a pharmacokinetic and bioequivalence study of amoxycillin/clavulanic acid formulations in calves.
Reyns
Tim
T
Ghent University, Faculty of Veterinary Medicine, Department of Pharmacology, Pharmacy and Toxicology, Salisburylaan 133, B-9820 Merelbeke, Belgium. Tim.Reyns@UGent.be
De Baere
Siegrid
S
Croubels
Siska
S
De Backer
Patrick
P
D000284:Administration, Oral; D000658:Amoxicillin / Q000097:blood; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D019818:Clavulanic Acid / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics; D012015:Reference Standards; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
D019818:Clavulanic Acid; D000658:Amoxicillin