Submitted to: Journal of Animal Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/22/2016
Publication Date: 12/29/2016
Citation: Smith, D.J., Shelver, W.L., Marx, A. 2016. Detection of residues in urine and tissues of sheep treated with trace levels of dietary ractopamine HCl. Journal of Animal Science. 94:5423–5433. doi:10.2527/jas2016-0899.
Interpretive Summary: Ractopamine HCl is a feed additive that is approved for use in cattle, swine, and turkey in the United States, Canada, and multiple other countries. Nevertheless, many countries to which the US exports meat products strictly prohibit the presence of ractopamine residues in food animal carcasses or edible offal. In addition, within the US, the presence of ractopamine in some specialty classes of livestock (show and competitive animals) is proscribed. Although there are many sensitive analytical methods capable of detecting trace levels of ractopamine, there are no published data demonstrating the minimal levels of ractopamine exposure that could cause animals to test positive for ractopamine. Thus it is unknown under what conditions accidental exposures or trace levels of ractopamine contamination in feeds could cause animals to test positive for ractopamine exposure. Our studies have indicated that ractopamine is detectable in feed and(or) urine of sheep exposed to trace levels of ractopamine (1/50th to 1/500Oth of the normal feed level). Measurement of ractopamine is dependent upon the matrix selected and the sensitivity of the assay used.
Technical Abstract: Ractopamine HCl was fed to sheep at 0 (Z), 0.001 (L), 0.01 (M), or 0.1 (H) mg/kg of diet (n = 4 per level, 0.5 kg of feed/day) for seven consecutive days and urine was collected daily about ~16 h post exposure. On-site lateral flow assays were able to reliably (0% false negatives) detect 20 ng of ractopamine HCl pre g of feed. Urine tested positive for ractopamine residues by lateral flow assay in 7.4 (Z), 0 (L), 82 (M), and 86% of the urine samples in each group. Using half maximum absorbance values as a cut off value, 0, 3.6, 86, and 93% of the Z, L, M, and H urine samples tested positive after ELISA analysis. Parent ractopamine was below the assay limit of quantification (0.7 ng/mL) in all urine samples using LC-MS/MS. After hydrolysis of ractopamine conjugates, total ractopamine (parent + hydrolyzed metabolites) in urine of L animals was always less than the LOQ, but in 7 of 28 samples were above the LOD (0.22 ng/mL); In contrast, urine in M animals contained 1.08-9.13 ng/mL of ractopamine, while urine of H animals contained 4.85-32.82 ng/mL of total ractopamine. Ractopamine is rapidly eliminated; nevertheless, urine from sheep exposed to as little as 5 µg/day (M) of ractopamine HCl had a greater than 80% chance of having a positive ractopamine urine sample detected by the screening assays which was confirmed by LC-MS/MS. Tissue residues of ractopamine were not detected in any of the sheep.