Determination of Flunixin in Edible Bovine Tissues Using Liquid Chromatography Coupled with Tandem Mass Spectrometry

Abstract

An accurate, reliable, and reproducible assay was developed and validated to determine flunixin in bovine liver, kidney, muscle, and fat. The overall recovery and percent coefficient of variation (%CV) of twenty-eight determinations in each tissue for flunixin free acid were 85.9% (5.9% CV) for liver, 94.6% (9.9% CV) for kidney, 87.4% (4.7% CV) for muscle, and 87.6% (4.4% CV) for fat. The theoretical limit of detection was 0.1 microg/kg (ppb, ng/g) for liver and kidney, and 0.2 ppb for muscle and fat. The theoretical limit of quantitation was 0.3, 0.2, 0.6, and 0.4 ppb for liver, kidney, muscle, and fat, respectively. The validated lower limit of quantitation was 1 ppb for edible tissues with the upper limit of 400 ppb for liver and kidney, 100 ppb for fat, and 40 ppb for muscle. Accuracy, precision, linearity, specificity, ruggedness, and storage stability were demonstrated. Briefly, the method involves an initial acid hydrolysis, followed by pH adjustment ( approximately 9.5) and partitioning with ethyl acetate. A portion of the ethyl acetate extract was purified by solid-phase extraction using a strong cation exchange cartridge. The eluate was then evaporated to dryness, reconstituted, and analyzed using LC/MS/MS. The validated method is sensitive and specific for flunixin in edible bovine tissue.

Excretory, Secretory, and Tissue Residues after Label and Extra-label Administration of Flunixin Meglumine to Saline- or Lipopolysaccharide-Exposed Dairy Cows

“…The FSIS Midwestern Laboratory used slight modifications of method CLG-FLX4.03. 20 Briefly, the method employed a Waters Acquity UPLC triple-quadrupole instrument with a 1.7 μm, BEH C18, 2.1 × 50 mm column and gradient mobile phase consisting of 0.4% formic acid (solvent A) and 4 parts of CH 3 Flunixin free acid in edible tissues, injection sites, and specialty tissues was determined at the USDA ARS Fargo laboratory using the method of Boner et al 17 on which method CLG-FLX4 20 is based. However, matrix-matched standard curves were prepared at 0.25, 1.25, 6.25, 12.5, and 18.75 ng flunixin free acid/mL of extract corresponding to tissue concentrations of 1, 5, 25, 50, and 75 ng/g, which allows quantitative analyses well below half the tolerance of skeletal muscle.…”

Determination of non-steroidal anti-inflammatory drugs residues in animal muscles by liquid chromatography–tandem mass spectrometry

“…Single-residue methods were published [4,5] but do not fulfil criteria of practicability. Recently, multi-residue methods for NSAIDs residues in milk [6] and kidneys [7] were published.…”

LC–MS/MS determination of the HIV-1 protease inhibitor indinavir in brain and testis of mice

“…the homogenization and extraction procedures, is critical. Due to the presence of solid cohesive (insoluble) material tissue samples have to be thoroughly destroyed, either mechanically [21][22][23][24] or chemically [25], in order to extract analytes. Then samples have to be clarified from tissue debris that may damage injection systems or analytical columns and purified from "soluble" tissue compounds (proteins or else) that may lead to apparatus pollution, unaccepted noise or interference with analytes.…”