Comparison of ELISA and LC-MS/MS for the measurement of flunixin plasma concentrations in beef cattle after intravenous and subcutaneous administration

Weilin L. Shelver, Lisa A Tell, Sarah Wagner, Scott E. Wetzlich, Ronald E. Baynes, Jim E. Riviere, David J. Smith

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Eight cattle (288 ± 22 kg) were treated with 2.2 mg/kg of body weight of flunixin free acid in a crossover design by subcutaneous (SC) and intravenous (IV) administration. After a minimum 1:10 dilution with 50 mM phosphate buffer, a commercial immunoassay was adapted to determine plasma concentrations of flunixin. The limit of detection was 0.42 ng/mL and the working range was 0.76-66.4 ng/mL when adjusted with the dilution factor. Plasma samples were extracted using mixed-mode cation exchange solid phase extraction prior to the LC-MS/MS analyses. The linear calibration curve for LC-MS/MS was 0.5-2000 ng/mL with a limit of detection of 0.1 ng/mL for flunixin and 0.3 ng/mL for 5-hydroxy flunixin. Flunixin concentrations determined using the ELISAs were compared to concentrations derived from the same samples using LC-MS/MS analyses. Pharmacokinetic parameters of time versus concentration data from each analysis were estimated and compared. Differences (P < 0.05) in estimates of area under the curve, volume of distribution, and clearance were apparent between ELISA and LC-MS/MS analyses after IV dosing; after SC dosing, however, there were no differences among the estimated parameters between the two methods. Quantitative immunoassay was a satisfactory method of flunixin analysis and that it would be difficult to differentiate routes of administration in healthy beef cattle based on the plasma elimination profile of flunixin after IV or SC administration.

Original languageEnglish (US)
Pages (from-to)2679-2686
Number of pages8
JournalJournal of Agricultural and Food Chemistry
Volume61
Issue number11
DOIs
StatePublished - Mar 20 2013

Fingerprint

flunixin
Beef
subcutaneous injection
intravenous injection
beef cattle
Intravenous Administration
Enzyme-Linked Immunosorbent Assay
enzyme-linked immunosorbent assay
Plasmas
immunoassays
Immunoassay
Dilution
Limit of Detection
detection limit
Pharmacokinetics
cation exchange
Solid Phase Extraction
solid phase extraction
tandem mass spectrometry
Cross-Over Studies

Keywords

  • administration
  • cattle
  • concentration
  • elimination
  • ELISA
  • flunixin
  • pharmacokinetics

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Chemistry(all)

Cite this

Comparison of ELISA and LC-MS/MS for the measurement of flunixin plasma concentrations in beef cattle after intravenous and subcutaneous administration. / Shelver, Weilin L.; Tell, Lisa A; Wagner, Sarah; Wetzlich, Scott E.; Baynes, Ronald E.; Riviere, Jim E.; Smith, David J.

In: Journal of Agricultural and Food Chemistry, Vol. 61, No. 11, 20.03.2013, p. 2679-2686.

Research output: Contribution to journalArticle

Shelver, Weilin L. ; Tell, Lisa A ; Wagner, Sarah ; Wetzlich, Scott E. ; Baynes, Ronald E. ; Riviere, Jim E. ; Smith, David J. / Comparison of ELISA and LC-MS/MS for the measurement of flunixin plasma concentrations in beef cattle after intravenous and subcutaneous administration. In: Journal of Agricultural and Food Chemistry. 2013 ; Vol. 61, No. 11. pp. 2679-2686.
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AU - Riviere, Jim E.

AU - Smith, David J.

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AB - Eight cattle (288 ± 22 kg) were treated with 2.2 mg/kg of body weight of flunixin free acid in a crossover design by subcutaneous (SC) and intravenous (IV) administration. After a minimum 1:10 dilution with 50 mM phosphate buffer, a commercial immunoassay was adapted to determine plasma concentrations of flunixin. The limit of detection was 0.42 ng/mL and the working range was 0.76-66.4 ng/mL when adjusted with the dilution factor. Plasma samples were extracted using mixed-mode cation exchange solid phase extraction prior to the LC-MS/MS analyses. The linear calibration curve for LC-MS/MS was 0.5-2000 ng/mL with a limit of detection of 0.1 ng/mL for flunixin and 0.3 ng/mL for 5-hydroxy flunixin. Flunixin concentrations determined using the ELISAs were compared to concentrations derived from the same samples using LC-MS/MS analyses. Pharmacokinetic parameters of time versus concentration data from each analysis were estimated and compared. Differences (P < 0.05) in estimates of area under the curve, volume of distribution, and clearance were apparent between ELISA and LC-MS/MS analyses after IV dosing; after SC dosing, however, there were no differences among the estimated parameters between the two methods. Quantitative immunoassay was a satisfactory method of flunixin analysis and that it would be difficult to differentiate routes of administration in healthy beef cattle based on the plasma elimination profile of flunixin after IV or SC administration.

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