A renewable screen-printed electrode based on magnetic NiCo@N-doped carbon nanotubes derived from Co-MOF for ractopamine detection.

A renewable electrochemical screen-printed electrode (SPE) is proposed based on magnetic bamboo-like nitrogen-carbon (N-C) nanotubes loaded with nickel-cobalt alloy (NiCo) nanoparticles (NiCo@N-CNTs) for the determination of ractopamine (RAC). During the preparation of NiCo@N-CNTs, Co-MOF-67 (ZIF-67) was firstly synthesized, and then blended with dicyandiamide and nickel acetate, followed by a one-step pyrolysis procedure to prepare NiCo@N-doped carbon nanotubes. The surface morphology, structure, and chemical composition of NiCo@N-CNTs were characterized by SEM, TEM, XRD, XPS, and EDS. The electrocatalytic and electrochemical behavior of NiCo@N-CNTs were investigated by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The results demonstrated that NiCo@N-CNTs possessed remarkable conductivity and electrocatalysis to the oxidation of ractopamine (RAC). By using screen-printed electrode (SPE), NiCo@N-CNTs, and a designed base support, a magnetic RAC sensor (NiCo@N-CNTs/SPE) was successfully constructed. It presented a detection linear range of 0.05-80 µM with a detection limit of 12 nM (S/N = 3). It also exhibited good sensitivity, reproducibility, and practicability in spiked real pork samples. Since the adhesion of NiCo/N-CNTs on SPE was controlled by magnet, the NiCo@N-CNTs was easily detached from the SPE surface by magnetism and thus displayed excellent renewability. This work broadened insights into portable devices for on-site and real-time analysis.

Helical au nanostructure for SERS detection of hazardous molecular and chiral enantiomers.

In recent years, incidents of pesticide pollution and abuse of feed additives have occurred frequently, which pose a great threat to human health. Raman spectroscopy has become an important method in the field of food safety due to its rapidity, simplicity and sensitivity. It is important to obtain complex structure to promote surface-enhanced Raman scattering (SERS) effect. In this study, gold helical nanoparticles with rich surface structure were synthesized using cysteine as induce agent. Notably, the complex helical structure and tip led to an excellent electromagnetic enhancement property. The helical structure showed ultra-sensitive detection of hazardous molecular, such as thiram and ractopamine. Interestingly, the D/L-Au structure had significant chiral optical activity and could be used as an unlabeled SERS platform for enantiomer identification. This study provided an effective strategy for the detection of pesticides and feed additives, which could be applied in other aspects of food safety in the future.

Cattle, carcass, economic, and estimated emission impacts of feeding finishing steers lubabegron or ractopamine hydrochloride.

Lubabegron (Experior; Elanco, Greenfield, IN, USA) is the first U.S. Food and Drug Administration approved feed additive for reducing gas emissions from feedlot animals or their waste; it does not have live or carcass performance claims. Our primary objective was to determine the effect of lubabegron on feedlot performance and carcass traits in finishing beef steers compared to ractopamine hydrochloride (Optaflexx; Elanco, Greenfield, IN, USA). A commercial feedlot trial using cross-bred beef steers (n = 2,117; 373 ±â€…15 kg initial body weight [BW]) was completed with a randomized complete block design. Treatments consisted of two feed additives: (1) OPT targeted to deliver 300 mg/animal/d of ractopamine hydrochloride for 28 ±â€…7 d out from harvest and (2) EXP targeted to deliver 36 mg/animal/d of lubabegron 56 ±â€…7 d out from harvest and a 4-d preslaughter withdrawal period. Twenty 70 to 142 hd pens with 10 pens per treatment were used. Cattle were weighed at arrival processing and at harvest and fed for an average of 167 d. Data were used to calculate production metrics, partial budgets, and estimated greenhouse gas emissions using published methods, and were analyzed using linear mixed models with pen as the experimental unit and block as a random intercept. A statistical significance threshold of α = 0.05 was determined a priori. There was no evidence for statistically significant differences between treatments for initial BW (P = 0.70), health-related outcomes (P values ≥ 0.43), or mobility scores (P = 0.09). Cattle-fed EXP had increased final BW, ADG, G:F, and decreased dry matter intake (P values ≤ 0.01) compared to OPT. Carcasses were 11 ±â€…1.76 kg (hot carcass weight) heavier in EXP group (P < 0.01), and differed between treatments for both yield grades (YG) and quality grades distributions (P values ≤ 0.01). Cattle-fed EXP had a shift toward more YG 1 and 2, select and sub-select carcasses compared to OPT, which had as shift toward more YG 3, 4, 5, prime and choice carcasses. With increased beef production and efficiency compared to OPT, the estimated CO2 equivalent emissions from production were reduced by 6.2% per unit of carcass weight for EXP (P ≤ 0.01). Estimated net returns/animal shipped were $56.61 ±â€…9.37 more for EXP than OPT (P ≤ 0.01). In conclusion, when cattle were fed for the same total number of days, feeding EXP compared to OPT increased net returns, feedlot performance, and efficiency, but resulted in carcass yield and quality characteristics that may impact marketing programs.

Immunological effects of ractopamine in rabbits receiving the viral inactivated rabbit hemorrhagic disease vaccine.

Background: There is an obvious lack of information about the effects of ractopamine, a ß-adrenergic agonist, on the growth performance and immune responses of rabbits, particularly in those receiving the viral rabbit hemorrhagic disease (RHD) vaccine. Aim: The current study was undertaken to study the effects of ractopamine on growth performances and immunological parameters in rabbits inoculated with the viral RHD vaccine. Methods: Experimental rabbits were grouped into four groups, the first acted as a control and received distilled water, the second received ractopamine, the third received inactivated RHD vaccine, and the fourth received both ractopamine, and inactivated RHD vaccine. Then, blood analysis, histopathological, histomorphometric, and immunohistochemistry (IHC) examinations were followed. Results: The obtained results demonstrated that ractopamine induced significant increases in body weight gain, neutrophils, monocytes, nitric oxide, lysosome, and improved feed conversion rate. A significant decrease in lymphocytes with insignificant decreases in eosinophils, phagocytic % and index, serum total protein, α, ß, and γ globulin were observed. Vaccinated rabbits only showed a marked rise in WBCs, neutrophils, monocytes, eosinophils, basophils, phagocytic index and activity, nitric oxide, lysosome activity, total protein, albumin, γ globulin, and a decrease in lymphocytes. Rabbits that received ractopamine and then vaccinated had insignificant increases in body weight, weight gain, WBCs, neutrophils, monocyte, eosinophils, basophils, phagocytic activity, and index, globulins besides a significant decrease in lymphocytes. Pathologically, rabbits that received ractopamine alone, with a vaccine or vaccinated only showed an increase in villus length, villus width, and absorption surface area. IHC of rabbits' liver and kidneys of the control and vaccinated group showed negative expression for caspase-3, but rabbits received ractopamine only or rabbits vaccinated and received ractopamine showed diffuse positive moderate expression for caspase-3. Conclusion: Ractopamine induced several adverse effects on the immune responses of the rabbits inoculated with the viral HRD vaccine.

Magnetic solid-phase extraction based on restricted-access molecularly imprinted polymers for ultrarapid determination of ractopamine residues from food samples by capillary electrophoresis.

An ultrafast, efficient, and eco-friendly method combining magnetic solid phase extraction and capillary electrophoresis with diode array detection have been developed to determine ractopamine residues in food samples. A restricted access material based on magnetic and mesoporous molecularly imprinted polymer has been properly synthesized and characterized, demonstrating excellent selectivity and high adsorbent capacity. Short-end injection capillary electrophoresis method was optimized: 75 mM triethylamine pH 7 as BGE, -20 kV, 50 mbar by hydrodynamic injection during 8 s, and capillary temperature at 25 °C; reaching ultrafast ractopamine analysis (∼0.6 min) with good peak asymmetry, and free from interfering and/or baseline noise. After sample preparation optimization, the conditions were: 1000 µL of sample at pH 6, 20 mg of adsorbent, stirring time of 120 s, 250 µL of ultrapure water as washing solvent, 1000 µL of methanol: acetic acid (7: 3, v/v) as eluent, and the adsorbent can be reused four times. In these conditions, the analytical method showed recoveries around to 100 %, linearity ranged from 9.74 to 974.0 µg kg-1, correlation coefficient (r) ≥ 0,99 in addition to adequate precision, accuracy, and robustness. After proper validation, the method was successfully applied in the analysis ractopamine residues in bovine milk and bovine and porcine muscle.

Comparison of immunoassay and LC-tandem mass spectrometry analyses of ractopamine in hog oral fluid.

The accurate detection of ractopamine in food animals is crucial for marketing since some entities require animals or animal carcasses to be free of ractopamine residues. Field-based ractopamine screening tests that are rapid, sensitive, and capable of high-throughput are highly desirable to ensure that inadvertent exposure to ractopamine did not occur in animals marketed as animals that have not been fed ractopamine. An immunochemically based lateral flow assay was used to analyze oral fluids from hogs never exposed to ractopamine and from hogs that were presumed positives and results were confirmed using an enhanced sensitivity LC-MSMS method. We found that an immunochemically based lateral flow system having a working range of 2.5 to 15 ng mL-1 worked well as a screening assay with 1.7% false positive results in freshly collected hog oral fluids. Using ractopamine glucuronide standards and LC-MSMS, we determined that the false positive results were not due to the presence of ractopamine glucuronide metabolites in oral fluids.

Core-Satellite Nanoassemblies as SPR/SERS Dual-Mode Plasmonic Sensors for Sensitively Detecting Ractopamine in Complex Media.

Highly sensitive and reliable detection of ß-adrenergic agonists is especially necessary due to the illegal abuse of growth-promoting feed additives. Here, we develop a novel surface plasmon resonance/surface-enhanced Raman scattering (SPR/SERS) dual-mode plasmonic sensor based on core-satellite nanoassemblies for the highly sensitive and reliable detection of ractopamine (RAC). The addition of RAC results in the decomposition of core-satellite nanoassemblies and consequently changes the Rayleigh scattering color of dark-field microscopy (DFM) images and the Raman scattering intensity of SERS spectra. The excellent sensitivity, specificity, and uniformity of this strategy were confirmed by detecting RAC in various complex media in the farm-to-table chain, and the limit of detection (LOD) was 0.03 ng/mL in an aqueous solution. In particular, the convenient access to livestock sewage not only ensures animal welfare but also provides great convenience for the market regulation of ß-agonists. The success of our on-site strategy only with a portable Raman device promises great application prospects for ß-agonist detection.

Feeding Ractopamine Improves the Growth Performance and Carcass Characteristics of the Lard-Type Mangalica Pig.

Mangalica pigs are gaining popularity within the U.S. as a niche breed, given their reputation for superior-quality pork. However, slow growth rates, a poor lean yield, and excessive adiposity limit the widespread adoption of Mangalica. To determine if feeding the metabolic modifier, ractopamine hydrochloride (RAC), would improve growth performance without impairing pork quality in the Mangalica, pigs were fed either 0 or 20 mg per kg RAC for 21 days. At 24 h postharvest, pork quality and carcass composition measurements were recorded; then, primal cuts were fabricated and assessed. RAC increased ADG (p < 0.04) and gain efficiency (p < 0.03) by 24% and 21%, respectively. RAC increased Loin Eye Area (p < 0.0001) by 21% but did not impact the 10th rib fat depth (p > 0.90) or marbling score (p > 0.77). RAC failed to alter any primal cut weights. Feeding RAC lowered b* values (p < 0.04) and tended to lower L* values (p < 0.08) while not affecting a* values (p > 0.30), suggesting RAC darkened loin color. Finally, RAC decreased cook yield percentage (p < 0.02) by 11% without impacting Warner-Bratzler Shear Force (p > 0.31). These data support the hypothesis that feeding RAC to Mangalica improves growth performance without impairing pork quality in this breed.

Beta-Adrenergic Agonists, Dietary Protein, and Rumen Bacterial Community Interactions in Beef Cattle: A Review.

Improving beef production efficiency, sustainability, and food security is crucial for meeting the growing global demand for beef while minimizing environmental impact, conserving resources, ensuring economic viability, and promoting animal welfare. Beta-adrenergic agonists and dietary protein have been critical factors in beef cattle production. Beta-agonists enhance growth, improve feed efficiency, and influence carcass composition, while dietary protein provides the necessary nutrients for muscle development and overall health. A balanced approach to their use and incorporation into cattle diets can lead to more efficient and sustainable beef production. However, microbiome technologies play an increasingly important role in beef cattle production, particularly by optimizing rumen fermentation, enhancing nutrient utilization, supporting gut health, and enhancing feed efficiency. Therefore, optimizing rumen fermentation, diet, and growth-promoting technologies has the potential to increase energy capture and improve performance. This review addresses the interactions among beta-adrenergic agonists, protein level and source, and the ruminal microbiome. By adopting innovative technologies, sustainable practices, and responsible management strategies, the beef industry can contribute to a more secure and sustainable food future. Continued research and development in this field can lead to innovative solutions that benefit both producers and the environment.

Magnetic and mesoporous molecularly imprinted polymer synthesized by rational computation design: Sample preparation and analysis of ractopamine.

Through density functional theory calculations was studied theoretically the formation process of a magnetic and mesoporous molecularly imprinted polymer for ractopamine (RAC), evaluating the molecular electrostatic potential map, functional monomers, functional monomer / template stoichiometry and crosslink agents. The results revealed that the best conditions for the synthesis were established with acrylic acid as functional monomer in a 1: 4 stoichiometry using acetonitrile as the solvent and ethylene glycol dimethacrylate as crosslink agent. It was observed that nine hydrogen bonds established between the RAC and acrylic acid play a key role on the pre-polymerization complex. In addition, three analytical methods using HPLC, UHPLC and CE instruments were optimized for rapid analysis. The adsorbent was experimentally synthesized considering the best conditions found at the molecular level and characterized by FTIR, DRX, TGA, SEM, TEM, surface analysis, and wettability. After that, the synthesized material was used in magnetic solid phase extraction combined with capillary electrophoresis in a preliminary RAC recovery study from milk samples. Finally, greenness metric with a score of 0.55 have been obtained for the sample preparation procedure using the online AGREEprep metric.

Nanoengineered disposable sensor fabricated with lanthanum stannate nanocrystallite for detecting animal feed additive: Ractopamine.

Ractopamine (RA) has been at the forefront of feed additives as a nutrient repartitioning mediator that recuperates the growth rate, decreases animal fat, and guarantees food safety. However, inappropriate and abusive usage of RA to enhance economic efficiency can negatively impact the environment-animal-human interactions. Therefore, the call for monitoring and quantifying RA is highly desired. In this work, the potentiality of La2Sn2O7 as an electrode modifier on the surface of the portable screen-printed carbon electrode (SPCE) was examined for its precision, disposability, and ability to detect RA. The superior electrocatalytic activity of the fabricated La2Sn2O7/SPCE fortifies its standpoints by displaying a wide linear working range of 0.01-501.2 µM, an enhanced sensitivity, a better stability, a lower LOD of 0.86 nM, and an increased selectivity toward the detection of RA. Furthermore, the investigation of the constructed electrochemical sensor with real-time food samples underpins its practicality and feasibility.

Glucometer-based biosensor for the determination of ractopamine in animal-derived foods using rolling circle amplification.

Screening for persistent organic pollutants (POPs) in food is a complex and challenging process, as POPs can be present in very low levels and can be difficult to detect. Herein, we developed an ultrasensitive biosensor based on a rolling circle amplification (RCA) platform using a glucometer to determine POP. The biosensor was constructed using gold nanoparticle probes modified with antibodies and dozens of primers, magnetic microparticle probes conjugated with haptens, and targets. After competition, RCA reactions are triggered, numerous RCA products hybridize with the ssDNA-invertase, and the target is successfully transformed into glucose. Using ractopamine as a model analyte, this strategy obtained a linear detection range of 0.038-5.00 ng mL-1 and a detection limit of 0.0158 ng mL-1, which was preliminarily verified by screening in real samples. Compared with conventional immunoassays, this biosensor utilizes the high efficiency of RCA and the portable properties of a glucometer, which effectively improves the sensitivity and simplifies the procedures using magnetic separation technology. Moreover, it has been successfully applied to ractopamine determination in animal-derived foods, revealing its potential as a promising tool for POP screening.

Au Nanoparticles Functionalized Covalent-Organic-Framework-Based Electrochemical Sensor for Sensitive Detection of Ractopamine.

Ractopamine, as a feed additive, has attracted much attention due to its excessive use, leading to the damage of the human nervous system and physiological function. Therefore, it is of great practical significance to establish a rapid and effective method for the detection of ractopamine in food. Electrochemical sensors served as a promising technique for efficiently sensing food contaminants due to their low cost, sensitive response and simple operation. In this study, an electrochemical sensor for ractopamine detection based on Au nanoparticles functionalized covalent organic frameworks (AuNPs@COFs) was constructed. The AuNPs@COF nanocomposite was synthesized by in situ reduction and was characterized by FTIR spectroscopy, transmission electron microscope and electrochemical methods. The electrochemical sensing performance of AuNPs@COF-modified glassy carbon electrode for ractopamine was investigated using the electrochemical method. The proposed sensor exhibited excellent sensing abilities towards ractopamine and was used for the detection of ractopamine in meat samples. The results showed that this method has high sensitivity and good reliability for the detection of ractopamine. The linear range was 1.2-1600 µmol/L, and the limit of detection (LOD) was 0.12 µmol/L. It is expected that the proposed AuNPs@COF nanocomposites hold great promise for food safety sensing and should be extended for application in other related fields.

Ractopamine residues in meat might reduce the risk of type 2 diabetes.

Background: The overall prevalence of diabetes in the world has risen substantially in the past several decades, so have complications and mortalities associated with it. Aim: Prevention strategies for diabetes thus become an urgent public health need for reducing the burden of diabetes. Methods: Ractopamine, a ß1/2-adrenergic receptor agonist, has been approved for use in finishing swine, cattle, and turkey in countries where meat exporting brings tremendous economic benefits. This leanness enhancer is recently found to be a full agonist at trace amine-associated receptor 1 also. A thorough literature review was performed to assess possible effects of ractopamine on glucose metabolism. Results: Activating ß-adrenoceptor could lead to glucose-lowering effects independent of insulin while activation on trace amine-associated receptor 1 induces an incretin-like signaling on insulin-secreting pancreatic ß-cells. Conclusion: Accordingly, it is hypothesized that long-term consuming meat containing ractopamine might lower the risk of type 2 diabetes.

Residues of ractopamine, a livestock feed additive, in meat might alleviate misuse of cocaine, nicotine, methamphetamine, and morphine.

Background: Substance misuse brings tremendous harm to global health. Strategies for the treatment and prevention of drug addiction are in urgent need. Aim: Trace amine-associated receptor 1 (TAAR1) widely distributed in the central nervous system has been identified as a hopeful target in the management of certain substance abuse. Discovery of food ingredients that act on TAAR1 might help health care providers develop chemoprevention for substance misuse disorders. Methods: Animal experiments clearly demonstrated the capability of TAAR1 agonists in attenuating addictive behavior regarding cocaine, nicotine, methamphetamine, and morphine. Ractopamine, a livestock feed additive used in the United States for over 20 years, has proven to be a full TAAR1 agonist. Literature review and internet web database survey were performed to see if ractopamine residues in meat could affect substance addiction behavior. Results: Integrating all available epidemiologic studies revealed that the prevalence of cocaine, nicotine, methamphetamine, and opioid misuse showed steadily downward or stable trends coincidently during the same time period of ractopamine use in the United States. Conclusion: A hypothesis is thus raised here that ractopamine residues in meat might have contributed secretly to the smoothened prevalence curves of cocaine, nicotine, methamphetamine, and opioids addiction.

Risk assessment supporting the establishment of a maximum residue limit for ractopamine in beef liver, applicable in the Arab Republic of Egypt.

In 2012, the Codex Alimentarius Commission adopted maximum residue limits (MRLs) for ractopamine in pig and cattle tissues. Egypt, a country that records a high consumption of beef liver, conducted a health risk assessment to estimate the risks associated with the adoption of Codex MRLs and the possible adoption of alternative values that may offer higher protection. Ractopamine was characterized based on previous assessments performed by international regulatory agencies, and an acceptable daily intake was set at 1 µg/kg bw for both chronic and acute ractopamine exposure. Beef liver consumption data for the Egyptian population were collected through a field survey (529 households, 1929 individuals). The standard body weight of 60 kg was used, as well as 70 kg, as a potentially more representative weight for the Egyptian population. Simulations showed that when the MRL for ractopamine in beef liver is set to 40 µg/kg (Codex MRL) or 20 µg/kg, the health-based guidance value of 1 µg/kg bw was not exceeded, as a result of chronic or acute exposure. An MRL of 20 µg/kg of ractopamine in beef liver was shown to provide optimum protection of Egyptian consumers, considering other potential sources of ractopamine intake and abnormally high consumption patterns, and was therefore recommended for adoption in Egypt. This study presents the inputs, model, and results of the probabilistic risk assessment that supported such recommendation. PRACTICAL APPLICATION: Residues of veterinary drugs, such as ractopamine, accumulate in animal tissues and may pose a risk to consumers. Establishing maximum residue limits (MRLs) will help importers by giving them the necessary visibility for commercial trade. It will also benefit Egyptian consumers, large consumers of beef liver, who will be better protected with a lower MRL than the internationally recommended one.

A new sensor for the rapid electrochemical detection of ractopamine in meats with high sensitivity.

In this research, g-C3N4/Cu@CoO/NC, which contained graphitic phase carbon nitride (g-C3N4) with a binary nanostructure and Cu@CoO/NC with a bimetallic MOF precursor, was constructed by a low-temperature pyrolysis process. The g-C3N4/Cu@CoO/NC was characterised by several techniques, including X-ray diffraction, scanning electron microscope, transmission electron microscope and X-ray photoelectron spectroscopy. Further, it was used to prepare an electrochemical sensor for the detection of ractopamine (RAC) in meat samples. The sensor showed excellent electrochemical oxidation characteristics for RAC detection, with a wide linear range (0.005 µmol/L to 32.73 µmol/L) and low detection limit (1.53 nmol/L). Meanwhile, the reproducibility, stability and interference of the g-C3N4/Cu@CoO/NC/GCE sensor were found to be excellent. Besides, the g-C3N4/Cu@CoO/NC/GCE sensor was well-used for the detection of RAC in pork, pig liver and lamb samples with recovery rates ranging from 96.5 % to 102.2 %.

Biochar-supported Cu nanocluster as an electrochemical ultrasensitive interface for ractopamine sensing.

Electrochemical sensors actually involve an electrocatalytic process involving an efficient and selective energy conversion that is related to the morphology and size of the interface of the modified materials. Ultrasmall nanoclusters or single atoms generate a greater catalytic ability than normal nanomaterials. In this study biochar-supported Cu nanoclusters (CuNCs@CNFs) were fabricated via a carbon confinement synthesis method toward ultrasensitive electrochemical sensing of ractopamine (RAC). RAC is a ß-adrenergic receptor agonist that is illegally used as a feed additive to significantly improve muscle accretion, resulting in RAC accumulation in meat-based food products. The unique structure of CuNCs@CNFs and the interconnectivity between the CuNCs and the CNFs enable the nanocomposite to significantly enhance conductivity and electrocatalytic activity. Using the CuNCs@CNFs-based sensor, RAC was determined with a high sensitivity of 1641 µA µM-1 cm-2. The feasibility of detecting RAC in spiked meat samples was also carried out with satisfactory recoveries ranging from 91.39 % to 94.58 %.

Ractopamine at the Center of Decades-Long Scientific and Legal Disputes: A Lesson on Benefits, Safety Issues, and Conflicts.

Ractopamine (RAC) is a synthetic phenethanolamine, ß-adrenergic agonist used as a feed additive to develop leanness and increase feed conversion efficiency in different farm animals. While RAC has been authorized as a feed additive for pigs and cattle in a limited number of countries, a great majority of jurisdictions, including the European Union (EU), China, Russia, and Taiwan, have banned its use on safety grounds. RAC has been under long scientific and political discussion as a controversial antibiotic as a feed additive. Here, we will present significant information on RAC regarding its application, detection methods, conflicts, and legal divisions that play a major role in controversial deadlock and why this issue warrants the attention of scientists, agriculturists, environmentalists, and health advocates. In this review, we highlight the potential toxicities of RAC on aquatic animals to emphasize scientific evidence and reports on the potentially harmful effects of RAC on the aquatic environment and human health.

Presence of ß2 -agonist growth promoters in human urine samples: GC-MS/MS evaluation of the excretion profiles of ractopamine administered in microdoses.

ß2 -adrenergic agonists having the potential to be misused to enhance performance for their thermogenic and anabolic properties are prohibited in sports. Clenbuterol, ractopamine and zilpaterol are utilised legally or illegally as growth promoters of animals raised for their meat. No withdrawal times are imposed for ractopamine prior to slaughter; residues are detected in meat of treated animals, which constitutes a risk of inadvertent consumption. Insufficient information is available on the fate of ractopamine in humans to implement efficient detection in athletes' urine samples. We have developed a confirmation procedure for total ractopamine in urine following the enzymatic hydrolysis of glucuronides and sulphates and the conversion to tri-TMS derivative (limit of identification at 0.15 ng/ml). The sulphates were found to form between 85% to 97% of ractopamine excreted in athletes' urine samples analysed routinely or in volunteers following the administration of a micro-dose of 2.5 µg. Peak levels were reached at 2 to 6 h and decreased rapidly below 1 ng/ml 10 h after dosing. With one exception, the highest level estimated in athletes' samples was 1.2 ng/ml. Zilpaterol was confirmed in a few urine samples collected in the USA and Mexico (highest level 2 ng/ml), while hundreds of athletes' samples were reported to contain clenbuterol by our laboratory over the past 7 years. Most of these cases originated from Mexico (n = 102) and Guatemala (n = 119), often clustered in events during which multiple samples were collected, and for the vast majority, in levels lower than 0.2 ng/ml.