A liquid chromatographic method is described for the determination of ractopamine hydrochloride (LY31537) in swine, turkey, and cattle feeds in the 1.25–100 ppm range and in swine supplement at 100 ppm. Feed samples were extracted with acidified methanol. An aliquot of the feed extract was diluted with water and buffered to pH 10.5 ± 0.5 with sodium carbonate to convert ractopamine hydrochloride to a free base. The free base was extracted from the buffered sample with ethyl acetate. The extract was further purified on an acid-washed, silica solid-phase extraction cartridge. After conversion of the free base back to the salt form with pH 4.5 buffer, analytical separation and quantitation of ractopamine hydrochloride were accomplished on IBM phenyl and Whatman ODS reversed-phase columns with coulometric detection at +600 mV. Mean daily recovery levels ranged from 85 to 100% for feeds fortified at 1.25 to 100 ppm. The coefficients of variation ranged from 1 to 6% for feeds fortified at 2.5 to 100 ppm.
Several enzyme immunoassay test kits are commercially available for screening of matrices including cattle feed, tissues, faeces and urine for the presence of beta-agonists. The kits utilized for this evaluation offer sensitivity as low as 0.1 ng ml-1 (assay solution concentration, not test sample). Evaluation of ractopamine hydrochloride (LY31537) solutions at concentrations as high as 1000 ng ml-1, using six different kits from four different manufacturers, showed cross-reactivity of less than 0.5%.
Experiments were conducted to determine the total residues remaining in ocular tissues of cattle and turkeys after oral administration of [14C]ractopamine HCl. Twelve cattle were intraruminally dosed with 0.9 mg x kg(-1) x d(-1) of [14C]ractopamine HCl for 7 d. Four cattle each were slaughtered with withdrawal periods of 48,96, and 144 h. Radioactive residues were not detectable in whole-eye homogenates from the cattle. Eight male and eight female turkeys per treatment received either 7.5, 22.5, or 30 ppm dietary [14C]ractopamine HCl (0.33, 1.02, and 1.36 mg x kg(-1) x d(-1); treatment groups 1, 2, and 3, respectively) for 7 d, and the birds were slaughtered with a 0-d withdrawal period. Eyes were dissected into retina/choroid/schlera (RCS), cornea/iris (CI), and aqueous humor (AH) fractions. Residues in RCS, CI, and AH of treatment 1 turkeys were not detectable. Residues in AH were < 0.02 ppm in treatment groups 2 and 3. Mean residues in RCS ranged from 0.15 to 0.26 ppm, and mean CI residues ranged from <0.09 to 0.17 ppm for treatment groups 2 and 3, respectively. The propensities of ractopamine and synthetic ractopamine metabolites to bind to melanin were studied in vitro using radiolabeled ligands with centrifugal filtration to separate melanin from unbound ligand. In vitro studies showed that [14C]ractopamine HCl binds to melanin rapidly and was displaced from melanin by other beta-agonists. Glucuronidation of ractopamine, which produced the major biotransformation product of ractopamine in all species studied to date, prevented binding to melanin. These studies demonstrate that the propensity for the in vivo binding of ractopamine HCl to pigmented ocular tissues is less than that reported for clenbuterol.
A liquid chromatographic (LC) method is described for determination of ractopamine hydrochloride (LY031537) in swine and turkey tissues. Liver, kidney, muscle, and fat samples were homogenized in methanol. An aliquot of the extract was evaporated, diluted with water, and buffered to pH 10.5 ± 0.5 with sodium carbonate to convert ractopamine hydrochloride to a free base. The free base was extracted from the buffered sample with ethyl acetate. The extract was further purified on a Bond Elut acid-washed silica solid-phase extraction cartridge. After converting the free base back to the salt form with pH 4.5 buffer, analytical separation and quantitation of ractopamine hydrochloride was performed on an IBM C18 column with coulometric detection at +600 mV. The limit of detection of the method was approximately 0.5 ppb as determined in swine liver. Overall recovery levels ranged between 75 and 100% for samples of liver, kidney, muscle, and fat fortified at 1 to 100 ppb. The coefficients of variation ranged from 2 to 18% for samples fortified at 1 to 100 ppb.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.