We tested whether periconceptional nutrition with cladodes from the cactus, Opuntia ficus-indica, with or without protein-enrichment, improved the metabolomic profile and reproductive outcomes of adult female sheep. Sixty Rambouillet ewes of similar body weight were randomly allocated among three nutritional treatments that were fed during the breeding period (34 days): Control (Control; n = 20), Opuntia (Opuntia; n = 20) and protein-enriched Opuntia (E-Opuntia; n = 20). There were no effects of treatment on body weight but assessment of urine samples indicated that, for 76 metabolites, the Control and Opuntia groups differed completely (P < 0.05), whereas there was overlap between the Control and E-Opuntia groups. It appears that, in Opuntia-fed and Control-fed sheep, different functional groups are activated leading to changes in the metabolism of glucose, tyrosine, methane, and glycerolipids. Fertility and reproductive rate tended to be higher in the Opuntia (70% and 95%) and E-Opuntia (90% and 110%) groups than in the Control (55% and 65%), and an orthogonal contrast revealed the difference between Control and Opuntia to be significant for both reproductive variables (P < 0.05). We conclude that nutritional supplementation with Opuntia cladodes, with or without protein enrichment, increased fertility rate and reproductive rate of female sheep, without any accompanying increases in body weight. Our observations suggest that the reproductive responses to Opuntia do not simply reflect a response to good nutrition, but might be caused by specific metabolites/metabolomic pathways, perhaps by an activation of the metabolism of glucose, methane, tyrosine and glycerolipids. There are few reports relating these metabolomic compounds with the metabolism of the sheep, let alone with reproductive efficiency. The novelty of these discoveries suggests that we need further research into the mechanisms through which nutrition affects the reproductive system.