The use of anabolic substances for growth promoting purposes in food producing animals is prohibited within the EU, yet ongoing applications of hormones such as oestradiol prove both difficult to detect and to distinguish from endogenous presence. Additionally, the misuse of glucocorticoid compounds (dexamethasone and prednisolone), which are permitted for therapeutic applications but can also promote improved animal health through long-term dosing, is reported to be increasing posing potential health concerns for consumers. Twenty-four male beef cattle were randomly assigned to four groups (n = 6) for experimental treatment over 40 days consisting of a control untreated group, and three treatment groups administered oestradiol, dexamethasone or prednisolone at levels known to reflect growth promoting practices. Untargeted metabolomic profiling of plasma collected from each animal midway through the study treatment period, was performed following reverse phase separation employing an UHPLC-QTof-MS system operating in positive electrospray ionization mode. Metabolomics analysis revealed metabolite perturbations in plasma common to all treated animals, with additional metabolites found to be specifically associated to the various differing growth promoting regimes. OPLS-DA modelling was used to discriminate plasma profiles of oestradiol, dexamethasone, or prednisolone from control untreated cohorts with 56, 48 and 58 ions found to be altered by the respective administered treatments. This culminated in 99 shared ions which could differentiate between plasma samples from untreated or variously growth promoter treated animals. Further assessment of these metabolites identified 24 ions to be significantly altered in comparison to control animals, of which 3, 11 and 8 ions were pertinent to oestradiol, dexamethasone or prednisolone administrations respectively and 2 relevant to more than one treatment. Incorporation of such markers, principally associated with lipid and fat metabolism responses to exogenous administrations, which are specific to growth promoting treatment types could be used in screening approaches to facilitate more effective confirmatory analysis.